3-aminopropan-1-ol

Endpoint summary

Description of key information

oral: In an oral two generation reproduction toxicity study (OECD TG 416 and GLP) in rats performed with the structural analogue substance 2-aminoethanol, the NOAEL for general toxicity was set at 300 mg/kg bw/day.

Obtained results from a metabolome study in rats show that 2-Aminoethanol and 3-Aminopropan-1-ol clustered well with each other in males and females and it could not be differentiated between the two metabolomic profiles.

inhalation: The exposure of rats to 2 -Aminoethanol, a structural analogue of 3 -Aminopropan-1-ol, for 28 days by inhalation caused concentration-related local lesions in larynx, trachea and lung. No systemic effects were observed. The NOAEC for systemic toxicity is the highest concentration of 150 mg/m3. The NOAEC for local effects is 10 mg/m3.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

Endpoint:

sub-chronic toxicity: oral

Remarks:

two-generation reproductive toxicity study

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 Aug 2006 - 15 Jan 2009

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 416 (Two-generation reproduction toxicity study)

Deviations:

yes

Remarks:

Food consumption was not determined between days 14 and 21 after parturition

Qualifier:

according to guideline

Guideline:

other: Corrigendum to EC Commission Directive 2004/73/EC, Part B: Methods for the determination of toxicity: Two-Generation Reproduction Toxicity Study; Official Journal of the European Comm unities; No. L216, pp. 236–246

Version / remarks:

29 Apr 2004

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.3800 (Reproduction and Fertility Effects)

Version / remarks:

Aug 1998

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: ad 1) JB116/2+3 (from 09 Aug – 04 Oct 2006); ad 2) JB116/4 (from 04 Oct – 29 Nov 2006); ad 3) JB116/9-17

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature, under N2

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI (Han)

Details on species / strain selection:

The rat is the preferred animal species for reproduction studies according to test guidelines. This strain was selected since extensive historical control data were available for Wistar rats.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services GmbH, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: (P) 44 (+/- 1) days
- Weight at study initiation: (P) Males: 162.1 (142.5 – 186.5) g; Females: 126.2 (110.6 – 145.1) g;
- Fasting period before study: none
- Housing: rats were housed individually in type DK III stainless steel wire mesh cages supplied by Be
cker & Co., Castrop-Rauxel, Germany (floor area of about 800 cm²), with the following exceptions:
• overnight mating: male and female mating partners were housed together in type DK III cages
• gestation day 18 – lactation day 21: pregnant animals and their litters were housed in Makrolon
type M III cages (floor area of about 800 cm²). The M III cages were also supplied by Becker & Co.
Pregnant females were provided with nesting material (cellulose wadding) toward the end of gestation.
- For enrichment wooden gnawing blocks (Typ NGM E-022, supplied by Abedd® Lab. and Vet. Service GmbH, Vienna, Austria) were added. The cages with the test animals were arranged in racks in such a way that uniform experimental conditions (ventilation and light) were ensured.
- Diet: ad libitum, ground Kliba maintenance diet mouse/rat “GLP” meal, supplied by Provimi Kliba SA,Kaiseraugst, Switzerland
- Water: ad libitum
- Acclimation period: 16 days
- Other: According to a written statement from the breeder, male and female animals were derived from different litters. This was necessary to rule out the possibility of sibling mating.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
The test substance (ethanolamine hydrochloride, EAH) was weighed and thoroughly mixed with a small amount of food. Then corresponding amounts of food, depending on the dose group, were added to this premix in order to obtain the desired concentrations. Mixing was carried out for about 10 minutes in a laboratory mixer. Test diets were prepared at intervals, which guaranteed that the test substance in the diet remained stable throughout the feeding period.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The stability of EAH in the diet over 32 days at room temperature was investigated analytically before the beginning of the study. Homogeneity and concentration control analyses were carried out at the beginning and toward the end of the premating periods. At least one analysis of test substance preparations for female animals was carried out during the gestation and lactation periods.

The analyses were carried out at the Analytical Chemistry Laboratory of Experimental Toxicology and Ecology of BASF SE, Ludwigshafen, Germany.

Duration of treatment / exposure:

semichronic duration (> 75 days)

Frequency of treatment:

continuously

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

300 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

25

Control animals:

yes, plain diet

Observations and examinations performed and frequency:

For parental animals:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily on working days and once daily on weekends

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes
- Time schedule for examinations: body weights of F0 and F1 parents were determined once weekly; during gestation and lactation F0 and F1 females were weighed on days 0, 7, 14 and 20 of gestation, and on days 1, 4, 7, 14 and 21 after birth.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Time schedule: once weekly (over a period of at least 6 days each) and weekly during gestation (days 0-7, 7-14, 14-20 post coitum; p.c.) and lactation periods (days 1-4, 4-7, 7-14 post partum; p.p.).

OTHER:
The F1 and F2 pups were sexed on the day of birth (day 0 p.p.) and weighed on days 1, 4, 7, 14, and 21 p.p. Their viability was recorded. At necropsy, all pups were examined macroscopically (including weight determinations of brain, spleen and thymus in one pup/sex/litter).

Serum concentrations of the test substance:
Blood samples were taken from all F0 and F1 parental animals of each sex and test group during week 10 of premating treatment and the plasma was analyzed for the concentration of Ethanolamine hydrochloride

Estrous cycle data were evaluated for F0 and F1 generation females over a three week period prior to mating until evidence of mating occurred. Moreover, the estrous stage of each female was determined on the day of scheduled sacrifice.

Parameters examined in [all/P/F1] male parental generations:
motility, sperm head count, morphology

Sacrifice and pathology:

For parental animals:
All F0 and F1 parental animals were sacrificed by decapitation under Isoflurane anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology, special attention was given to the reproductive organs. The liver, kidneys, adrenal glands, testes, epididmides. Cauda epididymis, prostate, seminal vesicles, ovaries, uterus, spleen, brain, pituitary gland and thyroid glands (with parathyroids) were weighed and the vagina, cervix uterie, uterus, ovaries, oviducts, left testis, left epididymis, seminal vesicles, coagulation glands, prostate, pituitary gland, adrenal glands, liver, kidneys, spleen, brain, thyroids (with parathyroids)and all gross lesions were fixed in an appropriate fixative, histologically processed and examined by light microscopy. From both ovaries (”ovary 1” and “ovary 2”) of F1 female animals (control and top dose), five sections were taken from the proximal and the distal part of the ovaries, at least 100 µm apart from the inner third of the ovary. All ovarian sections were prepared and evaluated for numbers of primordial and growing follicles.
As soon as possible after termination, one portion of the liver (lobus medialis) of each 10 dams per group was sampled to be analyzed for choline concentration.

Statistics:

See below

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

F0: No clinical signs or changes of the general behavior, which may be attributed to the test substance, were detected in F0 male or F0 female parental animals of the test groups 01 and 02 (100 and 300 mg/kg bw/d). Intensively yellow discolored urine was recorded in all F0 males and females of test group 03 (1000 mg/kg bw/d) from study week 3 onwards until the end of the treatment period. This urine discoloration mirrored the systemic availability of the test substance rather than being an adverse effect and was most likely caused by the excreted test compound and/or its metabolites.
Clinical observations for females during gestation of F1 litters:
All F0 females of test group 03 showed intensively yellow discolored urine during the entire gestation period for F1 litter. No other clinical findings were observed in the test groups 00-03 (0, 100, 300 and 1000 mg/kg bw/d). One sperm positive female of test group 02 (300 mg/kg bw/day) and one of test group 03 (1000 mg/kg bw/day) did not deliver F1 pups. This observation was not considered to be associated to the test compound.
Clinical observations for females during lactation of F1 litters:
All F0 females of test group 03 showed intensively yellow discolored urine during the entire lactation period for F1 litter. One high-dose female (1000 mg/kg bw/d) had just one pup (female), which was cannibalized by its mother on lactation day 8. No other clinical findings were observed in the test groups 00-03 (0, 100, 300 and 1000 mg/kg bw/day).
F1: No clinical signs or changes in general behavior, which may be attributed to the test substance, were detected in F1 male or F1 female parental animals of the test groups 11 and 12 (100 and 300 mg/kg bw/day).
Intensively yellow discolored urine was recorded in all F1 males and F1 females of test group 13 (1000 mg/kg bw/day) from study week 0 onwards until the end of the treatment period. This urine discoloration mirrored the systemic availability of the test substance rather than being an adverse effect and was most likely caused by excreted test compound and/or its metabolites. Furthermore, one F1 male animal of test group 11 (100 mg/kg bw/d) had a skin lesion at its throat during study weeks 3-6.
Clinical observations for females during gestation of F2 litters:
All F1 females of test group 13 showed intensively yellow discolored urine during the entire gestation period (F2 litter). No other clinical findings were observed in the test groups 10-13 (0, 100, 300 and 1000 mg/kg bw/day). One sperm-positive female of test group 10 (control), one of test group 11 (100 mg/kg bw/day) and two of test group 13 (1000 mg/kg bw/day) did not deliver F2 pups. These observations were not considered to be associated to the test compound due to a missing dose-response relationship.
Clinical observations for females during lactation of F2 litters: All F1 females of test group 13 showed intensive yellow discolored urine during the entire lactation period for F2 litters. No other clinical findings were observed in the test groups 10-13 (0, 100, 300
and 1000 mg/kg bw/day).

Mortality:

no mortality observed

Description (incidence):

There were no unscheduled mortalities of male and female parental animals in any test group.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

F0: Mean body weights and average body weight gain of the F0 parental males of all test substance-treated groups (100, 300 and 1000 mg/kg bw/d) were comparable to the controls throughout the entire study period. Observed differences between controls and test groups were regarded as spontaneous in nature. During premating, gestation and lactation periods, the mean body weights and body weight gains of the F0 parental females in the low- and mid-dose groups were generally comparable to the
concurrent control group. During premating, gestation and lactation periods, the mean body weights and body weight gains of the F0 parental females in the low- and mid-dose groups were generally comparable to the concurrent control group. Mean body weights and body weight gains of the F0 females in test group 03 (1000 mg/kg bw/d) were similar to the controls throughout the entire premating period. During gestation, these animals gained less weight from gestation day 7 onwards (up to 38%). As a consequence, body weights on gestation day 20 were 8% lower than the control. This effect may have been caused by the statistically significantly increased postimplantation loss and the statistically significantly decreased mean number of delivered pups in test group 03. This is also
indicated by the unaffected body weight of the high dose dams on post-delivery day 1. Mean body weights of the high-dose females remained comparable to the controls during entire lactation, w hereas the weight gain wavered up and down in the individual lactation sections.
F1: Mean body weights and body weight gain of the F1 parental males in test groups 11-13 (100, 300 and 1000 mg/kg bw/day) were comparable to the control throughout the entire treatment period. The statistically significantly decreased values of body weight gain in the high-dose males during study weeks 6-7 and 9-10 were in the normal range of fluctuation of this group and the control during the course of the study and, therefore, regarded as incidental. Mean body weights and body weight gain of the F1 parental females in test groups 11-12 (100 and 300 1000 mg/kg bw/d) were comparable to the control throughout premating, gestation and lactation periods. Mean body weights and body weight gains of the F1 females in test group 03 (1000 mg/kg bw/d) were similar to the controls throughout the entire premating period, the statistically significantly increased body weight gain of the high-dose F1 females (1000 mg/kg bw/d) during premating week 1-2 was regarded as incidental variance. The average weight gain of these animals was significantly below control (26%) during gestation days 14-20, which led to an averaged decrease of weight gain for the entire gestation of 17%. This effect may have been caused by the statistically significantly increased postimplantation loss and the statistically significantly decreased mean number of delivered pups in test group 03. Mean body weights of the high-dose females remained comparable to the controls during entire lactation.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

F0: Food consumption of the F0 parental males of all test substance-treated groups was generally comparable to the controls throughout the entire study. The statistically significant increase of food consumption in test group 02 (300 mg/kg bw/d) during study week 12-13 was regarded as spontaneous in nature. Food consumption of the F0 parental females of the low- and mid-dose groups (100 and 300 mg/kg bw/day) was comparable to the control animals during the periods of premating, gestation and lactation. Food consumption of the high-dose F0 females (1000 mg/kg bw/day) was also comparable during premating and gestation periods. However, in these animals, food consumption was statistically significantly below controls (up to 18%) in the individual lactation sections: days 1-4, 4-7, and 7-14 p.p.
F1: Food consumption of F1 male and female animals in test groups 11-12 (100, and 300 mg/kg bw/day) was generally comparable to the control group throughout the entire treatment period, covering premating, gestation and lactation periods. Food consumption of the high-dose F0 females (1000 mg/kg bw/day) was also comparable during premating and gestation periods. However, in these animals, food consumption was statistically significantly below controls (-11%) on lactation days 1-4, and rem ained below control on lactation days 4-7 and 7-14 (-7%), although not statistically significant.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

In the top-dose F0 and F1 males the test substance administration led to a decrease of absolute and relative organ weights of cauda epididymidis and epididymides. Furthermore, prostate weight and the number of homogenization resistant caudal epididymal sperm was slightly, but significantly decreased in the F0 males. These findings were considered to be treatment-related effects, whereas histomorphological correlates were missing.
A statistically significant increase of absolute and relative kidney weights was noted in male and female F1 animals of the mid (300 mg/kg bw/day) and top-dose (1000 mg/kg bw/day) groups. Because no histomorphological correlate was detected, the treatment-related weight increase was considered to be of no toxicological concern. As compared to control animals, the kidneys of low-,mid-, and top-dose male and female animals revealed a low incidence of basophilic tubules in a slightly higher number of animals. The severity (minimal to slight) was comparable between controls and treated animals and a clear dose-response relationship was missing. Thus this finding was considered to have no toxicological relevance.

Gross pathological findings:

no effects observed

Description (incidence and severity):

F0: All gross lesions observed in test animals occurred singularly. They are considered to be spontaneous lesions in origin and are not related to treatment. The female animal which was not pregnant as
well as the male mating partner did not show relevant gross lesions.
F1: All gross lesions observed in test animals occurred singularly. They are considered to be spontaneous lesions in origin and are not related to treatment. One non-pregnant female animal did show a bilaterally severe reduced size of the ovaries as well as a moderate thickening of the uterus wall. The other three non-pregnant females did not show any gross lesions. The four male mating partners did not show any gross lesions either.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

- Extramedullar hematopoiesis was seen in the spleen of 1000 mg/kg bw/day male animals in a slightly higher number of animals compared to control animals, but only in a minimal (grade 1) to slight (grade 2) severity. A treatment-related increase seems unlikely, because no weight deviations were observed, the severity was very low and comparable to the control animals. All other findings noted were single observations either, or were similarly in distribution pattern and severity in control rats compared to treatment groups. All of them are consideredto be incidental and/or spontaneous in origin and without any relation to treatment.
- Fertility: The non-pregnant female and the male mating partner did not show histopathological findings explaining the infertility. Only a minimal focal atrophy of the prostate was present, which is considered an incidental finding.
F1:
- As compared to control animals, the kidneys of low, mid, and top dose male and female animals revealed a low incidence of basophilic tubules in a slightly higher number of animals. The severity (minimal to slight) was comparable between controls and treated animals and a clear dose-response relationship was missing. The cauda epididymis and epididymides of top dose males showed no histomorphological correlates to the decreased organ weights. All other findings noted were single observations either, or were similarly in distribution pattern and severity in control rats compared to treatment groups. All of them are considered to be incidental and/or spontaneous in origin and without any relation to treatment.
- Fertility: One non-pregnant female showed a bilateral moderate diffuse stromal hyperplasia and a unilateral severe focally extensive chronic inflammation of the ovaries as well as an ovarian cyst. There were still corpora lutea present and the histopathological findings did not correlate with the gross lesion. The gross lesion “thickening of uterine wall” had no corresponding histological finding. The findings on the ovaries might explain the infertility. The male mating partner did not reveal lesions affecting the fertility. One female Animal showed within the uterus a severe multifocal endometrial and glandular degeneration and within the oviducts a severe diffuse epithelial degeneration, which explains the infertility of this animal. The male mating partner did not reveal lesions affecting the fertility. The other two mating pairs did not show any lesions affecting the fertility.

Histopathological findings: neoplastic:

no effects observed

Key result

Dose descriptor:

NOAEL

Remarks:

parental

Effect level:

300 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

food consumption and compound intake

organ weights and organ / body weight ratios

Key result

Critical effects observed:

no

Test substance stability:

The stability of test substance in rat diet was demonstrated for a period of 32 days at room temperature in a different batch of comparable quality, which was not used for the study. The homogeneity of the mixtures was verified. The concentration control analyses of the samples taken revealed that the values were within a range of 90-110% of the nominal concentration in all analyses at all time points, with the exception of one concentration in the feed of the high-dose group (88%).

Plasma concentrations of 2 -aminoethanol were below 3 mg/kg for all control animals, <3 - 4 mg/kg for the low dose animals, 8 - 11 mg/kg for the mid dose animals and 60 – 81 mg/kg for the high dose animals.

Toxicokinetic dataof 2 -aminoethanol (calculated as 2 -aminoethanol hydrochloride)fromthis two-generation reproduction toxicity studyshow a dose dependency of the plasma levels of 2 -aminoethanol in the experimental animals and there with prove the bioavailability of 2 -aminoethanol hydrochloride in principle.

 

Table 1 Mean test substance intake (mg/kg bw/d; minimum value / maximum value)

	Test group 01 (100 mg/kg bw/day)	Test group 02 (300 mg/kg bw/day)	Test group 03 (1000 mg/kg bw/day)
F0 males	94.3 (72.4 / 102.5)	283.2 (218.4 / 309.4)	943.3 (716.7 / 1032.6)
F0 females (premating)	96.7 (80.5 / 100.7)	289.6 (241.2 / 304.9)	964.4 (792.4 / 1017.8)
F0 females (F1 litter) - gestation period - lactation period*	103.5 (92.6 / 111.6) 99.2 (81.6 / 120.2)	315.2 (284.8 / 337.9) 306.7 (249.7 / 370.3)	1043.2 (989.4 / 1084.7) 866.0 (668.6 / 1053.9)

* = Days 1–14 p.p. only

Table 2 Absolute organ weights (P-generation)

Compared to the controls (= 100%), the following values (in %) were significantly changed (printed in bold):

	Male animals			Female animals
Group	01 100 mg/kg bw/d	02 300 mg/kg bw/d	03 1000 mg/kg bw/d	01 100 mg/kg bw/d	02 300 mg/kg bw/d	03 1000 mg/kg bw/d
Brain	99%	100%	97%*
Cauda epididymis	99%	102%	88%**
Epididymides	100%	101%	92%**
Prostate	92%	99%	86%**
Spleen				105%*	107%	97%

 *: p≤0.05; **: p≤0.01

All other mean absolute weight parameters did not show significant differences compared to the control groups.

The decrease of absolute weights of cauda epididymis, epididymides, and prostate in male top-dose animals (1000 mg/kg bw/d) were considered as treatment-related effects.

The decrease of brain weights in top-dose males (1000 mg/kg bw/d) as well as the increase of spleen weights in low-dose females (100 mg/kg bw/d) was considered as incidental and not treatment-related due to a missing dose-response relationship.

Table 3 Absolute organ weights (F1 generation)

Compared to the controls (= 100%), the following values (in %)were significantly changed (printed in bold):

 

	Male animals			Female animals
Group	11 100 mg/kg bw/day	12 300 mg/kg bw/day	13 1000 mg/kg bw day	11 100 mg/kg bw/day	12 300 mg/kg bw/day	13 1000 mg/kg bw/day
Cauda epididymis	96%	99%	88%**
Epididymides	100%	101%	91%**
Kidneys	99%	106%*	111%**	103%	106%**	115%**
Spleen	99%	103%	92%*
Thyroid glands	106%	99%	109%*	110%	118%**	111%*

 *: p≤0.05; **: p≤0.01

All other mean absolute weight parameters did not show significant differences compared to the control groups.

The decrease of absolute weights of cauda epididymis and epididymides in male top-dose animals (1000 mg/kg bw/d) were considered to be treatment-related.

The increase of absolute kidney weights of male and female animals in mid- (300 mg/kg bw/d) and top-dose (1000 mg/kg bw/d) groups, respectively, was statistically significant. Because no histomorphological correlate was detected, a treatment-related weight increase was less likely.

The decrease of spleen weights in top-dose males as well as the increase of thyroid glands in top-dose males and mid- and top-dose females, respectively, is considered incidental and not treatment-related due to a missing dose-response relationship.

Reference 2

Endpoint:

sub-chronic toxicity: oral

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

See read-across justification attached in chapter 13.

Reason / purpose for cross-reference:

read-across source

Key result

Dose descriptor:

NOAEL

Effect level:

369 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

food consumption and compound intake

organ weights and organ / body weight ratios

Remarks on result:

other: Correction for molecular weight applied (1.23)

Key result

Critical effects observed:

no

Reference 3

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Qualifier:

no guideline followed

Principles of method if other than guideline:

no data

GLP compliance:

not specified

Species:

rat

Strain:

not specified

Sex:

not specified

Details on test animals or test system and environmental conditions:

no data

Route of administration:

oral: feed

Vehicle:

not specified

Details on oral exposure:

no data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

6 weeks

Frequency of treatment:

daily

Dose / conc.:

860 mg/kg diet

No. of animals per sex per dose:

no data

Control animals:

not specified

Details on study design:

Post-exposure period: no data

Positive control:

no data

Observations and examinations performed and frequency:

no data

Sacrifice and pathology:

no data

Other examinations:

no data

Statistics:

no data

Clinical signs:

no effects observed

Mortality:

not specified

Body weight and weight changes:

not specified

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 specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

According to the authors there were no signs of toxicity. No further information available.

Dose descriptor:

NOAEL

Effect level:

860 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

not specified

Critical effects observed:

not specified

Reference 4

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

no guideline followed

Principles of method if other than guideline:

The plasma metabolite profile (metabolome) of rats administered via diet with the test substance 3-Aminopropan-1-ol or 2-Aminoethanol for 28 days was investigated. The metabolome as investigated in this study refers to 274 endogenous metabolites in plasma.
The analysis of the metabolome was performed by means of GC/MS and LC/MSMS in an unbiased broad profiling approach. The toxicological interpretation of the metabolome data is based on the information contained in the database MetaMap® Tox.

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 70 ± 1 days

Route of administration:

oral: feed

Vehicle:

not specified

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

28 days

Frequency of treatment:

daily

Dose / conc.:

6 000 ppm

Remarks:

3-Aminopropan-1-ol

Dose / conc.:

15 000 ppm

Remarks:

3-Aminopropan-1-ol

Dose / conc.:

3 000 ppm

Remarks:

2-Aminoethanol

Dose / conc.:

6 000 ppm

Remarks:

2-Aminoethanol

No. of animals per sex per dose:

5

Control animals:

yes

Positive control:

data set contained positive contol values

Observations and examinations performed and frequency:

All animals were checked daily for mortality and clinical signs.
Food consumption was determined on study days 7, 14, 21 and 28.
Body weight was determined before the start of the administration period in order to randomize the animals and on study days 0, 4, 7, 14, 21 and 28.

Other examinations:

METABOLITE PROFILING:
Blood samples for metabolome analysis were taken by puncturing the retrobulbar venous plexus on study day 7, 14 and 28 from overnight fasted animals under isoflurane anesthesia and the obtained EDTA-plasma was covered with nitrogen and frozen at -80°C. GC-MS (gas chromatography-mass spectrometry) and LC-MS/MS (liquid chromatography-MS/MS) were used for broad profiling, as described in van Ravenzwaay et al. (2007). Absolute quantification was performed by means of stable isotope-labelled standards. For all metabolites, changes were calculated as the ratio of the mean of metabolite levels in individual rats in a treatment group relative to mean of metabolite levels in rats in a matched control group (time point, dose level, sex).

Statistics:

The sex- and day-stratified heteroscedastic t-test ("Welch test") was applied to compare metabolite levels of dose groups with respective controls (see "Any other information on materials and methods").

Clinical signs:

no effects observed

Mortality:

not specified

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weight was not significantly altered neither in males nor in females.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food consumption was not significantly altered neither in males nor in females.

Details on results:

CHANGED METABOLITES - ASSESSMENT OF KEY CHANGES
15000 ppm 3-Aminopropan-1-ol
- induced slight changes in the metabolite profile compared to the control in male animals (no. of significantly changed metabolites about 2 to 3 times above the false positive rate at p < 0.05 of 14 metabolites; based on 274 possible metabolites). The effect of the treatment with 3-Aminopropan-1-ol on female animals was slightly stronger (no. of significantly changed metabolites about 4 to 5 times above the false positive rate at p < 0.05 of 14 metabolites; based on 274 possible metabolites). The profile strength is in good accordance with the findings of absolute metabolite changes also indicating a slightly stronger effect on female animals.
- in male animals, plasmalogen levels were significantly decreased. In females this was also the case, but here also sphingosines, fatty acid and cholesterol levels were significantly decreased
6000 ppm 3-Aminopropan-1-ol
- induced a slightly stronger effect in females, at least at day 28 of the treatment with metabolite changes lying up to 4 times higher than the range of the false positive rate at p < 0.05 of 14 metabolites. Overall, the test substance had a slightly weaker effect on the metabolome compared to the high dose of the substance. The profile strength is in good accordance with the findings of absolute metabolite changes also indicating a slightly stronger effect on female animals.
- in male animals the levels of two polyunsaturated fatty acids were downregulated as well as spermidine levels. In females cholesterol levels were significantly downregulated
6000 ppm (2-Aminoethanol)
- induced slight changes in the metabolite profile compared to the control in male and female animals (no. of significantly changed metabolites about 2 to 3 times above the false positive rate at p < 0.05 of 14 metabolites; based on 274 possible metabolites). The profile strength is in good accordance with the findings of absolute metabolite changes also indicating a similar effect on male and female animals, which is slightly stronger in female animals.
- in male animals, amino acids levels were significantly increased. In females this was not found, but here especially cholesterol was significantly decreased as well as glucuronic acid levels
3000 ppm (2-Aminoethanol)
- induced a slightly stronger effect in females, however lying more or less in the range of the false positive rate at p < 0.05 of 14 metabolites. In males and females the test substance had a weaker effect on the metabolome compared to the high dose of the substance. The profile strength is in good accordance with the findings of absolute metabolite changes also indicating a slightly stronger effect on female animals.
- in male animals only 18-Hydroxy-11-deoxycorticosterone levels were upregulated. In females no metabolites were consistently regulated at both sampling time points.
PATTERN RANKING
15000 ppm (3-Aminopropan-1-ol)
The comparison of the metabolite changes induced by 3-Aminopropan-1-ol treatment against the established specific metabolite patterns present in MetaMap® Tox (pattern ranking) did yield no matches for male animals. Female animals treated with 3-Aminopropan-1-ol showed a weak match with the pattern for estrogenic effects.
6000 ppm (3-Aminopropan-1-ol)
The comparison of the metabolite changes induced by the treatment with 3-Aminopropan-1-ol against the established specific metabolite patterns present in MetaMap® Tox (pattern ranking) did not yield any matches or weak matches in male animals. In female animals a weak match for estrogenic effects was obtained after 28 days of treatment.
6000 ppm (2-Aminoethanol)
The comparison of the metabolite changes induced by 2-Aminoethanol treatment against the established specific metabolite patterns present in MetaMap® Tox (pattern ranking) did yield a weak match with liver toxicity as typically induced by fibrates, phthalates and phenoxy herbizides after 28 days of treatment in male animals only. Female animals treated with 2-Aminoethanol showed no matches with the existing patterns.
3000 ppm (2-Aminoethanol)
The comparison of the metabolite changes induced by the treatment with 2-Aminoethanol against the established specific metabolite patterns present in MetaMap® Tox (pattern ranking) did not yield any matches or weak matches.
PROFILE COMPARISON WITH REFERENCE COMPOUNDS
15000 ppm (3-Aminopropan-1-ol)
Using total profile comparison, the metabolite profile of the high dose of 3-Aminopropan-1-ol in male animals did match well with the metabolic changes induced by 2-Aminoethanol and the low dose of 3-Aminopropan-1-ol. In total the profile comparison resulted in 19 matches with the best match being obtained for the high dose of 2-Aminoethanol. The other matches did not indicate a common mode of action. Using total profile comparison, the metabolite profile of the high dose of 3-Aminopropan-1-ol in female animals did match well with the metabolic changes induced by its low dose and 2-Aminoethanol. In total the profile comparison resulted in 9 matches with the best match being obtained for the low dose of 3-Aminopropan-1-ol.
6000 ppm (3-Aminopropan-1-ol)
Using total profile comparison, the metabolite profile of the low dose of 3-Aminopropan-1-ol in male animals matched only with the metabolic changes induced by the high dose of 3- Aminopropan-1-ol. However, the high dose treatment of 2-Aminoethanol was only slightly below the threshold. In female animals the total profile comparison resulted in 3 matches with the best match being obtained for the low dose of 2-Aminoethanol.
6000 ppm (2-Aminoethanol)
Using total profile comparison, the metabolite profile of the high dose of 2-Aminoethanol in male animals did match well with the metabolic changes induced by 3-Aminopropan-1-ol. In total the profile comparison resulted in 24 matches with the best match being obtained for the high dose of 3-Aminopropan-1-ol. The other matches did not indicate a common mode of action. Using total profile comparison, the metabolite profile of the high dose of 2-Aminoethanol in
female animals did match well with the metabolic changes induced by 3-Aminopropan-1-ol and the low dose of 2-Aminoethanol. In total the profile comparison resulted in 3 matches with the best match being obtained for the low dose of 3-Aminopropan-1-ol.
3000 ppm (2-Aminoethanol)
Using total profile comparison, the metabolite profile of the high dose of 2-Aminoethanol in male animals did only result in one match, the metabolic changes induced by 3-Aminopropan-1-ol. In female animals the total profile comparison resulted in 3 matches with the best match being obtained for the low dose of 3-Aminopropan-1-ol.
COMPARISON OF 2-AMINOETHANOL; 3-AMINOPROPAN-1-OL AND DIETHANOLAMINE
The metabolite profiles gained by treatment with 2-Aminoethanol, 3-Aminopropan-1-ol and the structurally similar Diethanolamine present in the MetaMapTox were compared to each other in a PCA (Principal Component Analysis). 2-Aminoethanol and 3-Aminopropan-1-ol clustered well with each other in males and females and it could not be differentiated between the two metabolomic profiles. Diethanolamine was clearly separated from 2-Aminoethanol and 3-Aminopropan-1-ol, which became more prominent with increasing dose.

To compare the similarity of the two treatments on a biological level a principal component analysis was conducted using the metabolic profiles emerging from the treatment with 2-Aminoethanol and 3-Aminopropan-1-ol. To further validate similarity, a third treatment with the structurally related compound Diethanolamine, was integrated as well, to determine which substance might serve as the best read across source for 3-Aminopropan-1-ol. Diethanolamine was clearly separated from 2-Aminoethanol and 3-Aminopropan-1-ol, which became more prominent with increasing dose, whereas the metabolic profiles of 2-Aminoethanol and 3-Aminopropan-1-ol were undistinguishable. Therefore, 2-Aminoethanol represented based on the systemic biological effects the better read across option for 3- Aminopropan-1-ol. In addition, the regulated metabolites of both treatments are presented. All comparisons show a very good overlap, except for the low dose in male animals. However, at a dose level of 3000 ppm 2-Aminoethanol effects only slightly above (5.6%) or even below (2.7%) the false discovery rate of 5% were detected. Regarding the differential resultsfor both substances obtained in the comparison against existing patterns for toxicological endpoints, an in-depth analysis for the estrogenic pattern as well as liver toxicity induced by phthalates, fibrates and phenoxy herbicides was conducted. Although for 2-Aminoethanol no match or weak match was obtained for the pattern for estrogenic effects, it was still ranked rather high. The relative effect of both substances relative to the reference substances used to create this pattern was estimated. This calculation is based on the values for each metabolite, which integrate not only significance but also the strength of regulation regarding each metabolite. By this approach both substances show a comparable effect at the same dose level (2-Aminoethanol HD and 3-Aminopropan-1-ol LD), which was however still considerably lower than for the used positive controls. The same was done for both treatments in regard of their effects on the liver. The strength of effect at comparable doses was slightly lower for 3-Aminopropan-1-ol (2-Aminoethanol HD and 3- Aminopropan-1-ol LD). However, both treatments showed a significantly lower effect compared to the used reference treatments.
Overall, the obtained results for 2-Aminoethanol and 3-Aminopropan-1-ol were highly comparable. These findings biologically support the use of 2-Aminoethanol as a source substance for the read-across of toxicological endpoints for 3-Aminopropan-1-ol.

Remarks on result:

not measured/tested

Critical effects observed:

not specified

Conclusions:

Based on the here obtained findings, Ethanolamine represents the best read across source for 3-Amino-1-propanol. The uncertainty provoked by the lack of repeated-dose toxicity data for 3-Aminopropan-1-ol could be reduced by an additional in vivo study comparing the metabolomic profile of rats treated either with Ethanolamine or 3-Aminopropan-1-ol, both chemicals providing “supporting evidence” for the applied read-across approach. These studies clearly demonstrate the metabolomic sameness of 3-Aminopropan-1-ol compared to Ethanolamine covering the endpoint repeated dose toxicity.

Executive summary:

In the present case-study a ´weight-of-evidence´ read-across approach from Ethanolamine (source) to 3 -Aminopropan-1 -ol (target) with metabolomics as ´supporting evidence´ reducing the remaining uncertainties is reported. We demonstrate the high structural similarity of the two analogous substances based on the available data and we report how metabolome data add confidence concerning mechanistic similarity in this read-across approach. Finally, the herein described read-across case supported by metabolomics is used to cover the data gaps in repeated dose and reproductive toxicity endpoint of 3 -Aminopropan-1 -ol via weight of evidence for the REACH-registration.

Based on the here obtained findings, Ethanolamine represents the best read across source for 3-Aminopropan-1 -ol. Slight effects on the liver are to be expected, which should be less severe for 3 -Aminopropan-1 -ol at a comparable dose level. A general impact of both treatments on lipid metabolism was observed, which is probably due to the natural occurrence of Ethanolamine and the structural similarity of 3-Aminopropan-1 -ol. Although the analysis revealed a hit for an estrogenic pattern, the underlying changes could be referred to a similar effect on liver parameters. As no changes in the hormone status itself could be detected, no proof for estrogenic activity of both treatments could be found in this study.

Reference 5

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

abstract

Qualifier:

no guideline followed

Principles of method if other than guideline:

Cats were treated with the test substance daily for 5 days/week for a total of 2 weeks.

GLP compliance:

no

Species:

cat

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 2.6-3.1 kg
- Diet ad libitum
- Water ad libitum

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

no data

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

2 weeks

Frequency of treatment:

5 times per week

Dose / conc.:

784 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

1 male and 2 female animals

Control animals:

not specified

Details on study design:

no data

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: before the study and on day 14 or on the day of death

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: No data
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals
- Parameters checked: no data

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: No data
- Animals fasted: No data
- How many animals: No data
- Parameters checked: No data

URINALYSIS: Yes
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, not specified
HISTOPATHOLOGY: Yes, not specified

Other examinations:

no data

Statistics:

no data

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

vomiting after administration of the substance, salivation and diarrhea.

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Details on results:

Three animals (2 females and 1 male) were treated 10-times within 2 weeks. All animals survived. No abnormalities were detected by gross pathology and histopathology after sacrifice of the animals. Clinical signs during the study included: vomiting after administration of the substance, salivation and diarrhea. The results of the examination of blood, urine, haemoglobin and liver function were not different from normal.

Dose descriptor:

NOAEL

Effect level:

ca. 780 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

gross pathology

Critical effects observed:

no

Reference 6

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

abstract

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rabbits were treated with the test substance daily for 5 days/week for a total of 2 weeks.

GLP compliance:

not specified

Species:

rabbit

Strain:

not specified

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Animal weight at study initiation: 2.40 - 2.65 kg
- Fasting period before study: none
- Food and water ad libitum

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

no data

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

2 weeks

Frequency of treatment:

5 times per week

Dose / conc.:

784 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

2 male and 1 female animal

Control animals:

no

Details on study design:

no data

Positive control:

not done

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: before the study and on day 14 or on the day of death

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: No data
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals
- Parameters checked: no data

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: No data
- Animals fasted: No data
- How many animals: No data
- Parameters checked: No data

URINALYSIS: Yes
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, not specified
HISTOPATHOLOGY: Yes, not specified

Other examinations:

no data

Statistics:

no data

Clinical signs:

no effects observed

Mortality:

mortality observed, non-treatment-related

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

effects observed, non-treatment-related

Description (incidence and severity):

The female animal had a slight diarrhea and the gross pathology after death showed a moderate pleuropneumonia and a pulmonary edema due to aspiration of the product.

Dose descriptor:

NOAEL

Effect level:

ca. 780 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Critical effects observed:

no

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

300 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

GLP study according OECD TG 416 (Read Across)

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)

Qualifier:

according to guideline

Guideline:

EU Method B.8 (Subacute Inhalation Toxicity: 28-Day Study)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: MEOA K 540
- Test substance No.: 08/0924-1

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: The stability under storage conditions over the exposure period was guaranteed by the manufacturer, and the manufacturer holds this responsibility.
- Stability under test conditions: Ambient

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI(Han)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH; Sandhofer Weg 7, 97633 Sulzfeld
- Age at study initiation: about 7 weeks
- Weight at study initiation: male ± 228 g; female ± 165 g
- Housing: The rats were housed together (up to 5 animals per cage) in Polysulfon cages (H-Temp [PSU]) (floor area about 2065 cm2). Bedding in the Polycarbonate cages were Type Lignocel fibres, dust free bedding. For enrichment wooden gnawing blocks (Typ NGM E-022), were added.
- Diet: mouse/rat laboratory diet “GLP”, 10 mm pellets (Provimi Kliba SA, Kaiseraugst, Basel Switzerland), ad libitum
- Water: tap water, ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: mixture of vapour and aerosol / mist

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 1.1 - <= 1.2 µm

Remarks on MMAD:

MMAD / GSD: The measurements of particle size in test group 3 resulted in MMADs of 1.1 and 1.2 µm with a GSD of 5.3 and 6.4.
The calculated mass fractions of particles below 3 µm aerodynamic size were 70.0 and 70.3 %. Thus the aerosols were highly respirable for rats and a very high proportion of the aerosol particles reached the lungs.

Details on inhalation exposure:

Generation of the inhalation atmospheres
Generator systems:
• Continuous infusion pumps PERFUSOR (B. Braun)
• Two-component atomizers (stainless steel, Schlick mod. 970)
Generation procedure:
The test substance was used unchanged.
For each concentration the test substance was supplied to a two-component atomizer at a constant rate by means of a metering pump. The aerosol was generated with compressed air mixed with conditioned dilution air into the inhalation system.
The control group was exposed to conditioned air.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentrations of the inhalation atmospheres were analysed by a gas chromatography analysis (GC) in all test groups including control. The vapor and liquid aerosol concentration were determined separately.
Daily means were calculated based on two measured samples per concentration and exposure. From the daily mean values of each concentration, mean concentrations and standard deviations for the entire study were derived.
In these groups, the constancy of concentrations in the inhalation systems in the chambers were continuously monitored using scattered light photometers.
In the control group (test group 0) one sample was analysed over the study period.

The particle size analysis was carried out with a cascade impactor. In test group 3, particle size distribution was determined two times during the exposure period. In this test group significant amount liquid aerosol was found and the concentration was high enough for this measurement. In test group 2, due to the low aerosol concentration, long sampling time was necessary. During ongoing sampling, deposited aerosol would get evaporated again and the measured particle size distribution would not reflect the real size distribution. Therefore, no cascade impactor measurement was performed in this test group. In test group 1, no significant aerosol fraction was determined.

Duration of treatment / exposure:

28 days

Frequency of treatment:

6 hours/day, 5 days/week

Dose / conc.:

10 mg/m³ air (nominal)

Dose / conc.:

50 mg/m³ air (nominal)

Dose / conc.:

100 mg/m³ air (nominal)

Dose / conc.:

10.2 mg/m³ air (analytical)

Remarks:

±2.7

Dose / conc.:

49.1 mg/m³ air (analytical)

Remarks:

±8.3

Dose / conc.:

155.9 mg/m³ air (analytical)

Remarks:

±23.4

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

The concentrations to be tested in this study were selected based on the results of the 5-day range finding study.
Summarizing the results, inhalation exposure to MEA for 6 hours per day on 5 consecutive days caused histological changes all over the respiratory tract. Most pronounced effects were observed in the upper respiratory tract.
At 500 mg/m3, minimal to mild inflammatory cell infiltrates in the submucosa of the ventral meatus in level I of the nasal cavity. In addition, four of the five animals revealed (multi)focal perivascular hemorrhage in this region. One animal showed necrosis of the squamous epithelium. In the area of the transition from squamous to the respiratory epithelium, four of the five animals revealed minimal to mild squamous metaplasia of the respiratory epithelium. In level II of the nasal cavity three of the five animals of the same concentration group showed minimal to moderate inflammatory cell infiltrates. In the larynx, minimal to severe epithelial necrosis, mild to severe inflammatory cell infiltrates, and minimal to moderate squamous metaplasia was observed. In level I of the larynx, inflammation was accompanied by necrosis of the submucosal glands. Moreover, cellular atypia within the metaplastic epithelium was observed in level I and II of the larynx. These findings were less severe in level III. Inflammatory cell infiltrates, focal epithelial necrosis and minimal diffuse epithelial hyperplasia could still be observed.
In the carina (trachea) respiratory epithelium hyperplasia and degeneration intermingled with inflammatory cell infiltrates in almost all animals of the high concentration group. In the lung minimal to mild hyperplasia of the bronchiolar epithelium in the areas of bifurcation of large bronchi was observed.
At 200 mg/m³ similar findings were noted in the above mentioned organs and tissues with less incidence and severity. At 20 mg/m3, no adverse effects were observed.
The observed effects seem to be associated with aerosol exposure. Considering the histological findings in the respiratory tract, 150 mg/m3 was selected as the high concentration for the main study to cause toxic effect. The mid concentration for the main study should be 50 mg/m3 because this concentration was around the saturated vapor concentration in the inhalation system. The low concentration should be 10 mg/m³, as the expected No Observed Adverse Effect Concentration (NOAEC).
150 mg/m³(61 ppm) as high concentration causing toxic effects
50 mg/m³ (20 ppm): as mid concentration
10 mg/m³ (4 ppm): as low concentration and expected NOAEC

Observations and examinations performed and frequency:

Mortality: The animals were examined for evident signs of toxicity or mortality twice a day (in the morning and in the late afternoon) on working days and once a day (in the morning) on Saturdays, Sundays and public holidays.

Clinical observations: The clinical condition of the test animals was recorded once during the pre-exposure period and on the post-exposure observation day and at least 3 times (before, during and after exposure) on exposure days. During exposure only a group wise examination was possible.

Body weight data: The body weight of the animals was determined at the start of the pre-exposure, at the start of the exposure period and then, as a rule, once a week as well as prior to gross necropsy. As a rule, the animals were weighed at the same time of the day. Body weight change was calculated as the difference between body weight on the respective exposure day and body weight on the day of the first exposure. Group means were derived
from the individual differences.

Food consumption: Food consumption was determined weekly and calculated as mean food consumption in grams per animal and day. The animals were maintained in social-housing cages, with 5 animals per cage, during the whole study period. Therefore, the food consumption was determined cage-wise. The food consumption per animal and day was calculated by dividing food consumption of the day of a respective cage by the 5 animals per cage. As the animals of each test group were housed in only one cage per sex, no statistical evaluation of food consumption is possible.

Ophthalmology: Before the start of the exposure period (day -3) the eyes of all animals, and at the end of the study (day 26) the eyes of all animals were examined for any changes in the refracting media with an ophthalmoscope (HEINE Optotechnik, Herrsching, FRG) after administration of a mydriatic (Mydrum, Chauvin ankerpharm GmbH, Rudolstadt, Germany).

CLINICAL PATHOLOGY
In the morning, blood was taken from the retro-orbital venous plexus from fasted animals. The animals were anaesthetized using isoflurane (Isoba®, Essex GmbH Munich, Germany). The blood sampling procedure and the subsequent analysis of the blood and serum samples were carried out in a randomized sequence. The assays of blood and serum parameters were performed under internal laboratory quality control conditions with commercial reference controls to assure reliable test results. The results of the clinical pathology examinations are expressed in units of the International System (SI). The following examinations were carried out in 5 animals per test group and sex.

Hematology: Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration (MCHC), Platelet count (PLT), Differential blood count, Reticulocytes, Prothrombin time.

Clinical chemistry: Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (ALP), γ-Glutamyltransferase (GGT), Sodium (NA), Potassium (K), Chloride (CL), Inorganic phosphate (INP), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Albumin (ALB), Globulins (GLOB), Triglycerides (TRIG), Cholesterol (CHOL), Magnesium (MG).

Sacrifice and pathology:

Necropsy: All animals were sacrificed under Narcoren anesthesia by exsanguination from the abdominal aorta and vena cava. The exsanguinated animals were necropsied and assessed by gross pathology.

Organ weights: The following weights were determined in all animals sacrificed on schedule:
1. Anesthetized animals
2. Adrenal glands
3. Brain
4. Epididymides
5. Heart
6. Kidneys
7. Liver
8. Lungs
9. Spleen
10. Testes
11. Thymus
12. Thyroid glands
3.10.3. Organ / Tissue fixation
The following organs or tissues were fixed in 4% buffered formaldehyde:
1. All gross lesions
2. Adrenal glands
3. Brain with olfactory bulb
4. Bone marrow (femur)
5. Eyes with optic nerve
6. Heart
7. Kidneys
8. Larynx/Pharynx
9. Liver
10. Lungs
11. Lymph nodes (tracheobronchial and mediastinal lymph nodes)
12. Nose (nasal cavity)
13. Esophagus
14. Ovaries
15. Seminal vesicle
16. Spinal cord (cervical, thoracic and lumbar cords)
17. Stomach (forestomach and glandular stomach)
18. Spleen
19. Testes
20. Thyroid glands
21. Thymus
22. Trachea
23. Urinary bladder
24. Uterus
From the liver, each one slices of the Lobus dexter medialis and the Lobus sinster lateralis were fixed in Carnoy’s solution and embedded in paraplast.

Histotechnical processing / Examination by light microscopy and assessment of findings: Fixation was followed by histotechnical processing and examination by light microscopy and assessment of findings according to the list below: Organs and tissues of main group animals designated for histological processing and light microscopic examination
1. All gross lesions
2. Nasal cavity (4 levels)
3. Larynx (3 levels)
4. Trachea
5. Lungs (5 lobes)
6. Lymph nodes (tracheobronchial and mediastinal lymph nodes)
7. Adrenal glands
8. Bone marrow (femur)
9. Brain
10. Heart
11. Kidneys
12. Liver
13. Esophagus
14. Ovaries
15. Seminal vesicles
16. Spinal cord (cervical, thoracic and lumbar cords)
17. Spleen
18. Stomach (forestomach and glandular stomach)
19. Testes
20. Thyroid glands
21. Thymus
22. Uterus
A correlation between gross lesions and histopathological findings was performed.

Statistics:

Body weight, body weight change: A comparison of each group with the control group was performed using DUNNETT's test (twosided) for the hypothesis of equal means.

Clinical pathology parameters, urine volume, urine specific gravity: Non-parametric one-way analysis using KRUSKALWALLIS test (two-sided).If the
resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using Wilcoxon-test (two-sided) for the equal medians.

Weight parameters: Non-parametric one-way analysis using KRUSKALWALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pair wise comparison of each concentration group with the control group was performed using the WILCOXON test for the hypothesis of equal medians.

Clinical signs:

no effects observed

Description (incidence and severity):

During the pre-exposure period and the post-exposure observation day the animals showed no clinical signs and findings different from normal. During the exposure period the animals of the control group showed no clinical signs and findings different from normal. During the exposure period a few animals crossbench all test groups showed salivation after exposure.

Mortality:

no mortality observed

Description (incidence):

No deaths were recorded throughout the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The mean body weight changes of the test substance exposed groups were not statistically significantly different from the control group 0.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No substance-related changes of food consumption were observed during the whole study period.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

no effects observed

Description (incidence and severity):

The ophthalmologic examinations did not show any impairment of the refracting media. Spontaneous findings such as remainders of the pupillary membrane or corneal stippling, striation of lens and opacity were observed in several animals of all test groups and the control group without any concentration-response relationship.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among hematological parameters were measured. In male rats of dose group 2 and 3 (50 mg/m3 and 150 mg/m3) the mean corpuscular hemoglobin concentration (MCHC) was higher compared to controls. The increase of this calculated parameter was not accompanied by an alteration of any other red blood cell parameter value. Therefore, the MCHC increase is regarded as possibly treatment-related, but not adverse.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related changes among clinical chemistry parameters were measured. At the end of the study, in male rats of all dose groups the creatinine values were higher compared to controls, whereas in females of dose group 1 (10 mg/m3) the urea levels were lower compared to controls. The values were not changed dose-dependently. Therefore, they are regarded as incidental and not treatment-related. In male rats of dose group 3 (150 mg/m3) the triglyceride values were decreased. This was the only altered clinical chemistry parameter and it was especially not accompanied by any change of protein, glucose or cholesterol levels. Therefore, this decreased triglyceride concentrations were regarded as not adverse (ECETOC Technical Report No. 85, 2002).

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

- Absolute weights: All mean absolute weight parameters did not show significant differences when compared with the control group 0.
- Relative organ weights: When compared with control group 0 (=100%), the mean relative weights of liver in male treatment groups were significantly decreased. All other mean relative weight parameters did not show significant differences when compared with the control group 0. The decrease of mean liver weights in treated males was not concentration dependent and there were no histopathological correlates. Therefore, the reduced liver weights in males of all treatment groups were regarded to be incidental and not related to treatment.

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no gross lesions in treated male and female animals.

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Histopathology Larynx: At the base of epiglottis (level I), a submucosal inflammation that was characterized by infiltrates of granulocytes and lymphoid cells occurred in all males and females of test groups 2 (50 mg/m3) and 3 (150 mg/m3). In animals of test group 3 (150 mg/m3), the inflammation was accompanied by degeneration of the submucosal glands. In addition, 4 males and 3 females of test group 3 (150 mg/m3) showed a focal epithelial necrosis at the base of epiglottis. In the same region, a focal squamous cell metaplasia was observed in 3 males and 2 females of test group 2 (50 mg/m3) as well as in all males and females of test group 3 (150 mg/m3). All these findings were related to treatment. The occurrence of a minimal inflammation at the base of epiglottis in one female of test group 1 (10 mg/m3) was considered incidental. A minimal or slight epithelial alteration was observed in 2 males and 3 females of the control group, in 4 males and one female of test group 1 (10 mg/m3), as well as in 2 males and 3 females of test group 2 (50 mg/m3). The epithelial alteration was located at the base of epiglottis and was characterized by a slight focal flattening of epithelial cells. The epithelial alteration was regarded as a spontaneous lesion. At the entrance to the ventral pouch (larynx, level II), a minimal (grade 1) focal squamous metaplasia was seen in one female of test group 2 (50 mg/m3) as well as in one male and two females of test group 3 (150 mg/m3). A minimal focal epithelial hyperplasia occurred in all males and in 4 females. A mostly minimal inflammation was observed in 2 males and 3 females of test group 2 (50 mg/m3) as well as in all males and 4 females of test group 3 (150 mg/m3). All findings were considered treatment-related.

Histopathology Trachea: In males, a minimal or slight focal squamous metaplasia that was located in the area of the carina occurred in 3 animals of test group 3 (150 mg/m3). A minimal or slight inflammation was observed in one male of test group 1 (10 mg/m3) and in 4 males of test group 3 (150 mg/m3). The occurrence of squamous metaplasia and of inflammation in males of test group 3 (150 mg/m3) was related to treatment. In females, a minimal focal inflammation was only seen in one control animal.

Histopathology Lungs: A minimal or slight focal or multifocal mucous cell hyperplasia was seen in single or few large bronchi in all males and 2 females of test group 3 (150 mg/m3). In affected bronchi, the number of goblet cells was minimally or slightly increased. The occurrence of mucous cell hyperplasia was regarded as treatment-related.

Histopathology rest: All other findings occurred either individually or were biologically equally distributed over the control group and the treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Histopathological findings: neoplastic:

not examined

Key result

Dose descriptor:

NOAEC

Remarks:

local effects

Effect level:

10 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

Key result

Dose descriptor:

NOAEC

Remarks:

systemic effects

Effect level:

150 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse systemic effect observed

Key result

Critical effects observed:

no

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

150 mg/m³

Study duration:

subacute

Species:

rat

Quality of whole database:

GLP study according OECD TG 412 (Read-Across)

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)

Qualifier:

according to guideline

Guideline:

EU Method B.8 (Subacute Inhalation Toxicity: 28-Day Study)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: MEOA K 540
- Test substance No.: 08/0924-1

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: The stability under storage conditions over the exposure period was guaranteed by the manufacturer, and the manufacturer holds this responsibility.
- Stability under test conditions: Ambient

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI(Han)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH; Sandhofer Weg 7, 97633 Sulzfeld
- Age at study initiation: about 7 weeks
- Weight at study initiation: male ± 228 g; female ± 165 g
- Housing: The rats were housed together (up to 5 animals per cage) in Polysulfon cages (H-Temp [PSU]) (floor area about 2065 cm2). Bedding in the Polycarbonate cages were Type Lignocel fibres, dust free bedding. For enrichment wooden gnawing blocks (Typ NGM E-022), were added.
- Diet: mouse/rat laboratory diet “GLP”, 10 mm pellets (Provimi Kliba SA, Kaiseraugst, Basel Switzerland), ad libitum
- Water: tap water, ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: mixture of vapour and aerosol / mist

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

>= 1.1 - <= 1.2 µm

Remarks on MMAD:

MMAD / GSD: The measurements of particle size in test group 3 resulted in MMADs of 1.1 and 1.2 µm with a GSD of 5.3 and 6.4.
The calculated mass fractions of particles below 3 µm aerodynamic size were 70.0 and 70.3 %. Thus the aerosols were highly respirable for rats and a very high proportion of the aerosol particles reached the lungs.

Details on inhalation exposure:

Generation of the inhalation atmospheres
Generator systems:
• Continuous infusion pumps PERFUSOR (B. Braun)
• Two-component atomizers (stainless steel, Schlick mod. 970)
Generation procedure:
The test substance was used unchanged.
For each concentration the test substance was supplied to a two-component atomizer at a constant rate by means of a metering pump. The aerosol was generated with compressed air mixed with conditioned dilution air into the inhalation system.
The control group was exposed to conditioned air.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentrations of the inhalation atmospheres were analysed by a gas chromatography analysis (GC) in all test groups including control. The vapor and liquid aerosol concentration were determined separately.
Daily means were calculated based on two measured samples per concentration and exposure. From the daily mean values of each concentration, mean concentrations and standard deviations for the entire study were derived.
In these groups, the constancy of concentrations in the inhalation systems in the chambers were continuously monitored using scattered light photometers.
In the control group (test group 0) one sample was analysed over the study period.

The particle size analysis was carried out with a cascade impactor. In test group 3, particle size distribution was determined two times during the exposure period. In this test group significant amount liquid aerosol was found and the concentration was high enough for this measurement. In test group 2, due to the low aerosol concentration, long sampling time was necessary. During ongoing sampling, deposited aerosol would get evaporated again and the measured particle size distribution would not reflect the real size distribution. Therefore, no cascade impactor measurement was performed in this test group. In test group 1, no significant aerosol fraction was determined.

Duration of treatment / exposure:

28 days

Frequency of treatment:

6 hours/day, 5 days/week

Dose / conc.:

10 mg/m³ air (nominal)

Dose / conc.:

50 mg/m³ air (nominal)

Dose / conc.:

100 mg/m³ air (nominal)

Dose / conc.:

10.2 mg/m³ air (analytical)

Remarks:

±2.7

Dose / conc.:

49.1 mg/m³ air (analytical)

Remarks:

±8.3

Dose / conc.:

155.9 mg/m³ air (analytical)

Remarks:

±23.4

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

The concentrations to be tested in this study were selected based on the results of the 5-day range finding study.
Summarizing the results, inhalation exposure to MEA for 6 hours per day on 5 consecutive days caused histological changes all over the respiratory tract. Most pronounced effects were observed in the upper respiratory tract.
At 500 mg/m3, minimal to mild inflammatory cell infiltrates in the submucosa of the ventral meatus in level I of the nasal cavity. In addition, four of the five animals revealed (multi)focal perivascular hemorrhage in this region. One animal showed necrosis of the squamous epithelium. In the area of the transition from squamous to the respiratory epithelium, four of the five animals revealed minimal to mild squamous metaplasia of the respiratory epithelium. In level II of the nasal cavity three of the five animals of the same concentration group showed minimal to moderate inflammatory cell infiltrates. In the larynx, minimal to severe epithelial necrosis, mild to severe inflammatory cell infiltrates, and minimal to moderate squamous metaplasia was observed. In level I of the larynx, inflammation was accompanied by necrosis of the submucosal glands. Moreover, cellular atypia within the metaplastic epithelium was observed in level I and II of the larynx. These findings were less severe in level III. Inflammatory cell infiltrates, focal epithelial necrosis and minimal diffuse epithelial hyperplasia could still be observed.
In the carina (trachea) respiratory epithelium hyperplasia and degeneration intermingled with inflammatory cell infiltrates in almost all animals of the high concentration group. In the lung minimal to mild hyperplasia of the bronchiolar epithelium in the areas of bifurcation of large bronchi was observed.
At 200 mg/m³ similar findings were noted in the above mentioned organs and tissues with less incidence and severity. At 20 mg/m3, no adverse effects were observed.
The observed effects seem to be associated with aerosol exposure. Considering the histological findings in the respiratory tract, 150 mg/m3 was selected as the high concentration for the main study to cause toxic effect. The mid concentration for the main study should be 50 mg/m3 because this concentration was around the saturated vapor concentration in the inhalation system. The low concentration should be 10 mg/m³, as the expected No Observed Adverse Effect Concentration (NOAEC).
150 mg/m³(61 ppm) as high concentration causing toxic effects
50 mg/m³ (20 ppm): as mid concentration
10 mg/m³ (4 ppm): as low concentration and expected NOAEC

Observations and examinations performed and frequency:

Mortality: The animals were examined for evident signs of toxicity or mortality twice a day (in the morning and in the late afternoon) on working days and once a day (in the morning) on Saturdays, Sundays and public holidays.

Clinical observations: The clinical condition of the test animals was recorded once during the pre-exposure period and on the post-exposure observation day and at least 3 times (before, during and after exposure) on exposure days. During exposure only a group wise examination was possible.

Body weight data: The body weight of the animals was determined at the start of the pre-exposure, at the start of the exposure period and then, as a rule, once a week as well as prior to gross necropsy. As a rule, the animals were weighed at the same time of the day. Body weight change was calculated as the difference between body weight on the respective exposure day and body weight on the day of the first exposure. Group means were derived
from the individual differences.

Food consumption: Food consumption was determined weekly and calculated as mean food consumption in grams per animal and day. The animals were maintained in social-housing cages, with 5 animals per cage, during the whole study period. Therefore, the food consumption was determined cage-wise. The food consumption per animal and day was calculated by dividing food consumption of the day of a respective cage by the 5 animals per cage. As the animals of each test group were housed in only one cage per sex, no statistical evaluation of food consumption is possible.

Ophthalmology: Before the start of the exposure period (day -3) the eyes of all animals, and at the end of the study (day 26) the eyes of all animals were examined for any changes in the refracting media with an ophthalmoscope (HEINE Optotechnik, Herrsching, FRG) after administration of a mydriatic (Mydrum, Chauvin ankerpharm GmbH, Rudolstadt, Germany).

CLINICAL PATHOLOGY
In the morning, blood was taken from the retro-orbital venous plexus from fasted animals. The animals were anaesthetized using isoflurane (Isoba®, Essex GmbH Munich, Germany). The blood sampling procedure and the subsequent analysis of the blood and serum samples were carried out in a randomized sequence. The assays of blood and serum parameters were performed under internal laboratory quality control conditions with commercial reference controls to assure reliable test results. The results of the clinical pathology examinations are expressed in units of the International System (SI). The following examinations were carried out in 5 animals per test group and sex.

Hematology: Leukocyte count (WBC), Erythrocyte count (RBC), Hemoglobin (HGB), Hematocrit (HCT), Mean corpuscular volume (MCV), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration (MCHC), Platelet count (PLT), Differential blood count, Reticulocytes, Prothrombin time.

Clinical chemistry: Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (ALP), γ-Glutamyltransferase (GGT), Sodium (NA), Potassium (K), Chloride (CL), Inorganic phosphate (INP), Calcium (CA), Urea (UREA), Creatinine (CREA), Glucose (GLUC), Total bilirubin (TBIL), Total protein (TPROT), Albumin (ALB), Globulins (GLOB), Triglycerides (TRIG), Cholesterol (CHOL), Magnesium (MG).

Sacrifice and pathology:

Necropsy: All animals were sacrificed under Narcoren anesthesia by exsanguination from the abdominal aorta and vena cava. The exsanguinated animals were necropsied and assessed by gross pathology.

Organ weights: The following weights were determined in all animals sacrificed on schedule:
1. Anesthetized animals
2. Adrenal glands
3. Brain
4. Epididymides
5. Heart
6. Kidneys
7. Liver
8. Lungs
9. Spleen
10. Testes
11. Thymus
12. Thyroid glands
3.10.3. Organ / Tissue fixation
The following organs or tissues were fixed in 4% buffered formaldehyde:
1. All gross lesions
2. Adrenal glands
3. Brain with olfactory bulb
4. Bone marrow (femur)
5. Eyes with optic nerve
6. Heart
7. Kidneys
8. Larynx/Pharynx
9. Liver
10. Lungs
11. Lymph nodes (tracheobronchial and mediastinal lymph nodes)
12. Nose (nasal cavity)
13. Esophagus
14. Ovaries
15. Seminal vesicle
16. Spinal cord (cervical, thoracic and lumbar cords)
17. Stomach (forestomach and glandular stomach)
18. Spleen
19. Testes
20. Thyroid glands
21. Thymus
22. Trachea
23. Urinary bladder
24. Uterus
From the liver, each one slices of the Lobus dexter medialis and the Lobus sinster lateralis were fixed in Carnoy’s solution and embedded in paraplast.

Histotechnical processing / Examination by light microscopy and assessment of findings: Fixation was followed by histotechnical processing and examination by light microscopy and assessment of findings according to the list below: Organs and tissues of main group animals designated for histological processing and light microscopic examination
1. All gross lesions
2. Nasal cavity (4 levels)
3. Larynx (3 levels)
4. Trachea
5. Lungs (5 lobes)
6. Lymph nodes (tracheobronchial and mediastinal lymph nodes)
7. Adrenal glands
8. Bone marrow (femur)
9. Brain
10. Heart
11. Kidneys
12. Liver
13. Esophagus
14. Ovaries
15. Seminal vesicles
16. Spinal cord (cervical, thoracic and lumbar cords)
17. Spleen
18. Stomach (forestomach and glandular stomach)
19. Testes
20. Thyroid glands
21. Thymus
22. Uterus
A correlation between gross lesions and histopathological findings was performed.

Statistics:

Body weight, body weight change: A comparison of each group with the control group was performed using DUNNETT's test (twosided) for the hypothesis of equal means.

Clinical pathology parameters, urine volume, urine specific gravity: Non-parametric one-way analysis using KRUSKALWALLIS test (two-sided).If the
resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using Wilcoxon-test (two-sided) for the equal medians.

Weight parameters: Non-parametric one-way analysis using KRUSKALWALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pair wise comparison of each concentration group with the control group was performed using the WILCOXON test for the hypothesis of equal medians.

Clinical signs:

no effects observed

Description (incidence and severity):

During the pre-exposure period and the post-exposure observation day the animals showed no clinical signs and findings different from normal. During the exposure period the animals of the control group showed no clinical signs and findings different from normal. During the exposure period a few animals crossbench all test groups showed salivation after exposure.

Mortality:

no mortality observed

Description (incidence):

No deaths were recorded throughout the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The mean body weight changes of the test substance exposed groups were not statistically significantly different from the control group 0.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No substance-related changes of food consumption were observed during the whole study period.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

no effects observed

Description (incidence and severity):

The ophthalmologic examinations did not show any impairment of the refracting media. Spontaneous findings such as remainders of the pupillary membrane or corneal stippling, striation of lens and opacity were observed in several animals of all test groups and the control group without any concentration-response relationship.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among hematological parameters were measured. In male rats of dose group 2 and 3 (50 mg/m3 and 150 mg/m3) the mean corpuscular hemoglobin concentration (MCHC) was higher compared to controls. The increase of this calculated parameter was not accompanied by an alteration of any other red blood cell parameter value. Therefore, the MCHC increase is regarded as possibly treatment-related, but not adverse.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related changes among clinical chemistry parameters were measured. At the end of the study, in male rats of all dose groups the creatinine values were higher compared to controls, whereas in females of dose group 1 (10 mg/m3) the urea levels were lower compared to controls. The values were not changed dose-dependently. Therefore, they are regarded as incidental and not treatment-related. In male rats of dose group 3 (150 mg/m3) the triglyceride values were decreased. This was the only altered clinical chemistry parameter and it was especially not accompanied by any change of protein, glucose or cholesterol levels. Therefore, this decreased triglyceride concentrations were regarded as not adverse (ECETOC Technical Report No. 85, 2002).

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

- Absolute weights: All mean absolute weight parameters did not show significant differences when compared with the control group 0.
- Relative organ weights: When compared with control group 0 (=100%), the mean relative weights of liver in male treatment groups were significantly decreased. All other mean relative weight parameters did not show significant differences when compared with the control group 0. The decrease of mean liver weights in treated males was not concentration dependent and there were no histopathological correlates. Therefore, the reduced liver weights in males of all treatment groups were regarded to be incidental and not related to treatment.

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no gross lesions in treated male and female animals.

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Histopathology Larynx: At the base of epiglottis (level I), a submucosal inflammation that was characterized by infiltrates of granulocytes and lymphoid cells occurred in all males and females of test groups 2 (50 mg/m3) and 3 (150 mg/m3). In animals of test group 3 (150 mg/m3), the inflammation was accompanied by degeneration of the submucosal glands. In addition, 4 males and 3 females of test group 3 (150 mg/m3) showed a focal epithelial necrosis at the base of epiglottis. In the same region, a focal squamous cell metaplasia was observed in 3 males and 2 females of test group 2 (50 mg/m3) as well as in all males and females of test group 3 (150 mg/m3). All these findings were related to treatment. The occurrence of a minimal inflammation at the base of epiglottis in one female of test group 1 (10 mg/m3) was considered incidental. A minimal or slight epithelial alteration was observed in 2 males and 3 females of the control group, in 4 males and one female of test group 1 (10 mg/m3), as well as in 2 males and 3 females of test group 2 (50 mg/m3). The epithelial alteration was located at the base of epiglottis and was characterized by a slight focal flattening of epithelial cells. The epithelial alteration was regarded as a spontaneous lesion. At the entrance to the ventral pouch (larynx, level II), a minimal (grade 1) focal squamous metaplasia was seen in one female of test group 2 (50 mg/m3) as well as in one male and two females of test group 3 (150 mg/m3). A minimal focal epithelial hyperplasia occurred in all males and in 4 females. A mostly minimal inflammation was observed in 2 males and 3 females of test group 2 (50 mg/m3) as well as in all males and 4 females of test group 3 (150 mg/m3). All findings were considered treatment-related.

Histopathology Trachea: In males, a minimal or slight focal squamous metaplasia that was located in the area of the carina occurred in 3 animals of test group 3 (150 mg/m3). A minimal or slight inflammation was observed in one male of test group 1 (10 mg/m3) and in 4 males of test group 3 (150 mg/m3). The occurrence of squamous metaplasia and of inflammation in males of test group 3 (150 mg/m3) was related to treatment. In females, a minimal focal inflammation was only seen in one control animal.

Histopathology Lungs: A minimal or slight focal or multifocal mucous cell hyperplasia was seen in single or few large bronchi in all males and 2 females of test group 3 (150 mg/m3). In affected bronchi, the number of goblet cells was minimally or slightly increased. The occurrence of mucous cell hyperplasia was regarded as treatment-related.

Histopathology rest: All other findings occurred either individually or were biologically equally distributed over the control group and the treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Histopathological findings: neoplastic:

not examined

Key result

Dose descriptor:

NOAEC

Remarks:

local effects

Effect level:

10 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

Key result

Dose descriptor:

NOAEC

Remarks:

systemic effects

Effect level:

150 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse systemic effect observed

Key result

Critical effects observed:

no

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

10 mg/m³

Study duration:

subacute

Species:

rat

Quality of whole database:

GLP study according OECD TG 412 (Read Across)

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

A sub-chronic toxicity study is not available for 3 -Aminopropan-1 -ol. However, a two-generation reproduction toxicity study performed with 2 -Aminoethanol, an analogue substance, is available. In addition, a subacute inhalation toxicity study is available for 2-Aminoethanol.

Oral

2 -Aminoethanol (MEA HCl), an analogue substance was tested in an oral two generation reproduction toxicity study according to OECD TG 416 under GLP-conditions (ACC and CEFIC, 2009). The test substance was administered to groups of 25 male and 25 female healthy young Wistar rats (F0 parental generation) as a homogeneous addition to the food in different concentrations, which were adjusted regularly to obtain target dose levels of 0, 100, 300 and 1000 mg/kg bw/day. At least 75 days after the beginning of treatment, F0 animals were mated to produce a litter (F1 generation). Regarding general repeated dose toxicity, the dose level of 1000 mg/kg bw/day caused systemic toxicity in parental females, as was indicated by reduced food consumption and/or body weight gain during gestation and lactation. In the mid and high dose F1 animals the absolute and relative kidney weights were statistical significantly increased without histopathological correlate findings. In the top-dose F0 and F1 males the test substance administration led to a decrease of absolute and relative organ weights of cauda epididymidis and epididymides. Furthermore, prostate weight and the number of homogenization resistant caudal epididymal sperm was slightly, but significantly, decreased in the F0 males. These findings were considered to be treatment-related effects, whereas histomorphological correlates were missing. Based on this study, the NOAEL for general toxicity was set at 300 mg/kg bw/day. Correction for molecular weight for the target substance resulted in a NOAEL of 369 mg/kg bw/day for 3-aminopropan-1-ol (factor 1.23).

A 14 -day subacute study was conducted with rabbits and cats which received 784 mg/kg/day 3-aminopropan-1-ol by gavage (BASF AG, 1968). The animal’s state of health was checked each day, examinations of blood, urine and clinical chemistry parameters were performed. Gross and histopathology were done at the end of the study period. No substance related effects were seen in the two weeks of the study. One female rabbit died probably due to aspiration of the product. The two male animals survived ten applications with slight reduction of body weights. No abnormalities were detected by gross pathology and histopathology of the 2 male animals and no clinical signs were observed during the study. The results of the examination of blood, urine, haemoglobin and liver function did not show any differences from normal.

In the study with cats all animals survived the 14 -day treatment procedure. No abnormalities were detected by gross pathology and histopathology after sacrifice of the animals (BASF AG, 1961). Clinical signs during the study included vomiting after administration of the substance, salivation and diarrhea. The results of the examination of blood, urine, haemoglobin and liver function were not different from normal. Excluding the death occurred accidently in the rabbit study the NOAEL for both species were defined at ca. 780 mg/kg bw/day based on the clinical findings and the results of pathology.

Another subacute feeding study was performed with rats at a dose level of 860 mg/kg bw/day (American Cyanamide Co, 1954). Here, rats were dosed for 6 weeks, but unfortunately no further details of the study are available. The authors set the NOAEL to 860 mg/kg bw/day.

The metabolite profile (metabolome) in plasma of rats treated with 3-Aminopropan-1-ol (15000 ppm group and 6000 ppm group) or 2-Aminoethanol (6000 ppm group and 3000 ppm group) for 28 day was investigated. Blood samples for metabolome analysis were taken by puncturing the retrobulbar venous plexus on study day 7, 14 and 28 from overnight fasted animals under isoflurane anesthesia. The metabolome as investigated in this study refers to 274 endogenous metabolites in plasma. The analysis of the metabolome was performed by means of GC/MS and LC/MS-MS in an unbiased broad profiling approach. The toxicological interpretation of the metabolome data is based on the information contained in the database MetaMap® Tox. More than 800 metabolome studies have been used to build the database "MetaMap®Tox". For all metabolites, changes were calculated as the ratio of the mean of metabolite levels in individual rats in a treatment group relative to mean of metabolite levels in rats in a matched control group.

The metabolite profiles gained by treatment with 3-Aminopropan-1-ol, 2-Aminoethanol and the structurally similar Diethanolamine present in the MetaMap®Tox were compared to each other in a Principal Component Analysis. 2-Aminoethanol and 3-Aminopropan-1-ol clustered well with each other in males and females and it could not be differentiated between the two metabolomic profiles. Diethanolamine was clearly separated from 2-Aminoethanol and 3-Aminopropanol-1-ol, which became more prominent with increasing dose.

The correlation analysis of the whole plasma metabolite profile of animals treated with 2-Aminoethanol against the metabolite profiles of the reference compounds in the MetaMap®Tox data base showed a very good correlation with 3-Aminopropan-1-ol with correlation coefficients up to 0.58 in males and 0.71 in females. In male animals a correlation coefficient above 0.4 and in female animals above 0.5 is indicating a significant correlation between two treatments. Correlation coefficients of 0.7 and higher are found in completely identical experiments using the same treatment. Especially in female animals, the metabolic changes induced by 2-Aminoethanol near-to-perfectly reflect the changes induced by 3-Amino-propan-1-ol. In all cases 3-Aminopropan-1-ol represented the best obtained match.

These findings biologically support the use of 2-Aminoethanol as a source substance for the read across of toxicological endpoints for 3-Aminopropan-1-ol. Slight effects on the liver are to be expected, which should be less severe for 3-Aminopropan-1-ol at a comparable dose level. A general impact of both treatments on lipid metabolism was observed, which is probably due to the natural occurrence of 2-Aminoethanol and the structural similarity of 3-Aminopropan-1-ol. Although the analysis revealed a hit for an estrogenic pattern, the underlying changes could be referred to a similar effect on liver parameters. As no changes in the hormone status itself could be detected, no proof for estrogenic activity of both treatments could be found in this study (BASF SE, 2017).

Inhalation

The toxicity of 2-Aminoethanol (MEA) was studied in a 28 days inhalation toxicity study performed according to OECD TG 412 under GLP-conditions (REACH Ethanolamines consortium, 2010). Groups of five male and five female Wistar rats per test group were exposed nose-only to dynamic atmosphere of MEA for 6 hours per day on 5 consecutive days per week for 4 weeks (28-day study). The target concentrations were 10 mg/m3, 50 mg/m3 and 150 mg/m3. A concurrent control group was exposed to conditioned air. Clinical observations, body weight determinations and food consumption determinations were performed for all animals. Ophthalmological examinations were performed prior to exposure and towards the end of the exposure. After the last exposure, blood was sampled from the animals and hematology and clinical chemistry parameters were determined. The animals were then subjected to gross necropsy (including macroscopic examination of the major internal organs and collection of organ weight data). Selected tissues were processed histopathologically and were evaluated by light microscopy. Histological examinations were performed according to standardized methods with particular emphasis on the nasal cavity (4 levels) and larynx (3 levels). The exposure of rats to MEA caused concentration-related lesions in larynx, trachea and lung. At the high concentration (150 mg/m3), submucosal inflammation (levels I, II) in males and females, degeneration of submucosal glands (level I) in males and females, focal epithelial necrosis (level I) in males and females, focal squamous metaplasia, (level I) in males and females; (level II) in one male and 2 females and focal epithelial hyperplasia (level II) in males and females were observed in the larynx. In the trachea, focal squamous metaplasia (carina) accompanied by inflammation in males was observed. At the mid concentration (50 mg/m3), submucosal inflammation (level I and II) in males and females and squamous metaplasia (level I and II) in few males and females in the larynx was reported. No treatment-related weight changes, gross lesions or microscopic findings at the low concentration (10 mg/m3). No histopathological effects were seen in any other organ outside the respiratory tract. The NOAEC for systemic toxicity is the highest concentration of 150 mg/m3. The NOAEC for local effect was the lowest tested concentration of 10 mg/m3 under the current test conditions.

Exposure based adaptation of information requirements:

According to REGULATION (EC) No 1907/2006, Annex VIII, IX and Annex X, repeated dose toxicity testing (section 8.6) may be omitted, if relevant human exposure can be excluded in accordance with Annex XI section 3. Furthermore and in accordance with section 3.2 (b) of Annex XI (as amended by Regulation 134/2009), testing for repeated dose toxicity can be omitted when the substance is not incorporated in an article and the manufacturer can demonstrate and document for all relevant scenarios that throughout the life cycle strictly controlled as well as rigorously contained conditions as set out in Article 18(4)(a) to (f) (Regulation 1907/2006) apply.

 

Life-cycle stage(s) covered:

 

1.      Manufacture of substance (PROCs 1, 2, 3,)

2.      Formulation of preparations (PROC 1, 2, 3)

3.      Charging and discharging of substances and mixtures (PROC 8b, 9)

4.      Use as intermediate (PROC 1, 2, 3)

5.      Use in gas treatment (PROC 1, 2, 3)

6.      Use in laboratories (PROC 15)

 

Classification:

Acute Tox. 4                      H302: Harmful if swallowed.

Skin Corr. 1B                     H314: Causes severe skin burns and eye damage.

Eye Dam. 1                        H318: Causes serious eye damage.

 

Data profile:

3-Aminopropanol (3-AP, CAS 156-87-6) is harmful after single uptake and is corrosive to the skin as well as the eyes. Based on a bacterial reverse mutation assay, a sister-chromatid exchange (SCE) assay, anin vitromammalian cell gene mutation test (HPRT), and anin vitromammalian cell micronucleus test the test substance is not considered to induce gene mutations neither in the absence nor in the presence of a metabolic activation system.

 

Manufacture of substance and formulation of preparation:

3-Aminopropanol (3-AP, CAS 156-87-6) is manufactured in a continuous multi-product plant by reaction of ethylene cyanohydrin with hydrogen in the presence of ammonia and a solid catalyst.

The feedstocks ethylene cyanohydrin, stored in large tanks, and hydrogen, sourced from the site hydrogen grid, are fed into a trickle bed reactor which is filled with a catalyst. In addition, Ammonia is added. The hydrogenation reaction operates at elevated pressure and temperature, with recirculation of hydrogen and ammonia gas and back-mixing of the reaction medium.

After cooling down, the reaction mixture, rich in 3-AP, is separated in gas and liquid by a two-pressure expansion. The gas phase is partially returned to hydrogenation, while the remaining liquid is sent to a distillation tower where residual ammonia is distilled off under pressure and recovered to be re-used in the synthesis step. At the bottom of the tower, crude 3-AP is discharged and pumped to dedicated intermediate storage tanks. As a measure of quality assurance, the crude product is analyzed twice a day. For sampling, workers wear stringent personal protective equipment (PPE), such as substance appropriate gloves, appropriate skin coverage as well as suitable eye protection (protective goggles and face shield). Thus, exposure is considered to be controlled and negligible.

The purification of crude 3-AP is carried out in campaigns in a continuous multi-product distillation train. 3-AP is transferred via pipeline from the intermediate storage tanks to a series of distillation towers where low and middle boilers as well as heavies are safely separated from the product. The finished product, withdrawn as the top product of the purification tower, is analyzed every few hours and pumped to one of two dedicated check tanks. As before, for sampling tasks workers wear stringent personal protective equipment (PPE), such as substance appropriate gloves, appropriate skin coverage as well as suitable eye protection (protective goggles and face shield). Thus, exposure is considered to be controlled and negligible.

When filled, the check tanks are sampled for quality control. If the material passes the quality and sensory testing, it will be transferred to the final 3-AP storage tank where tank trucks and rail cars are loaded for delivery to internal and external customers. Loading takes place at a dedicated facility specifically designed and operated for the transfer of hazardous substances. In addition to the technical risk management measures in place, workers wear stringent PPE. Thus, exposure is considered to be controlled and negligible.

The production plants as well as all infrastructure as to steam, condensate, raw materials, products and wastes are designed to avoid open handling of materials and thus the exposure of operators, technicians and mechanics to 3-AP is controlled and negligible.

Both production lines and corresponding tank farms are well equipped with operational and monitoring devices to control and adjust process parameters (pressure, temperature, liquid level, flows, etc.). Safety instrumented systems are in place to prevent these parameters from reaching unsafe levels that could result in unacceptable risks to employees or the environment.

Product contamination due to product changeovers are obviated in both synthesis and distillation train by flushing all pieces of equipment with methanol and/or water depending on the solubility behavior of the previous product. The flushing media are sent via pipeline to incineration for disposal.

While producing 3-AP, only little waste water is generated which requires effluent treatment in the waste water treatment plant on site. Liquid waste streams resulting mainly from the distillation process are of organic nature and are collected in buffer tanks, until they are disposed of via the existing disposal systems for light and heavy boiling residues (pipeline and tank truck deliveries to residue incineration plant on site).

Exhaust piping from the synthesis and distillation train is connected to an incinerator of an adjacent production unit. To avoid air emissions of hazardous materials securely, the exhaust gas will be directed automatically to an emergency flare, should the incinerator fail.

Potential hazards for operators, technicians and mechanics, arising from any activities or tasks (e.g. sampling, loading/unloading of tank truck and rail cars, leak repair etc.) related to production assets are evaluated and documented in risk assessments and work permits. Safety procedures and exposure mitigation measures (e.g. personal protective equipment to wear) are in place.

 

Use as intermediate:

3-Aminopropanol (3-AP, CAS 156-87-6) is used as intermediate under strictly controlled conditions as a starting material for the synthesis of D-panthenol.Potential hazards for operators, technicians and mechanics, arising from any activities or tasks (e.g. sampling, loading/unloading of tank truck and rail cars, leak repair etc.) related to production assets are evaluated and documented in risk assessments and work permits. Safety procedures and exposure mitigation measures (e.g. personal protective equipment to wear) are in place.

 

Use in Gas Treatment and Charging and Discharging:

3-Aminopropanol (3-AP, CAS 156-87-6), in combination with other amines, is applied as an aqueous solution in closed-loop processes for capturing CO₂from gas streams, mainly flue gas.

The gas stream containing CO₂is introduced through an exhaust pipe into the purification plant. The exhaust gas in the exhaust pipe is introduced into a direct contact cooler where the exhaust gas is washed and humidified by counter-current flow to water. The cooled and humidified exhaust gas is then introduced into an absorber, where the exhaust gas is brought in counter-current flow to an aqueous absorbent solution (containing 3-AP and other amines) introduced through a lean amine pipe. Rich absorbent having absorbed CO₂is collected at the bottom of the absorber and is withdrawn trough a rich absorbent pipe, whereas the CO₂lean exhaust gas is released through a lean exhaust pipe after being washed in washing sections by means of water recycled through washing water cooling circuits. Because of extensive emission control systems and the relatively low volatility of 3-AP, the emission with treated flue gas will be very low and exposure to the public will be neglectable.

The rich absorbent is heated against the lean absorbent by means of a heat exchanger before being introduced into a regeneration column where the rich absorbent is stripped by counter-current flow to steam. The stripping steam is generated in a reboiler in which the lean absorbent collected at the bottom of the regeneration column is introduced through a lean absorbent withdrawal pipe.

Lean absorbent is withdrawn from the reboiler through the lean absorbent pipe and recycled into the absorber. Steam and CO₂liberated from the absorbent in the regeneration column is washed in washing sections by counter-current flow to water recirculating in washing water cooling circuits, before being withdrawn through a CO₂collection pipe. The CO₂and steam are cooled in a condenser and condensed water is removed in a separator and recycled into the regeneration column through a recycling line. The partly dried CO₂is withdrawn through a pipe for further treatment and downstream use.

The purification plant is equipped with sampling ports for gas and liquid, temperature and pressure probes and measurements of gas/liquid flow in all parts of the plant.

Waste products are incinerated, and waste water if any is routed to waste water treatment. Waste products are handled as hazardous waste by an approved waste handling company according to local regulations. Dedicated procedures are in place for all operating steps involving the substance handling, e.g. procedures for transfer of process inventory, draining of process units, clean-up of spillage, waste handling, etc.

Storage equipment such as the amine make-up tank for the process is sealed or nitrogen blanketed. The make-up tank could be charged a few times year from a road tanker/bulk transport with very limited exposure of workers due to the above described precautions. Accidental spills of 3-AP to the environment are avoided, as the purification plant is placed in a bunded area.

In view of the high degree of automation and closed nature of this process, exposure is limited to sampling procedures, cleaning and maintenance of equipment. Risk management measures are constantly applied by the highly trained and restricted personnel to minimize exposure.

 

Rigorous containment measures:

3-Aminopropanol (3-AP, CAS 156-87-6) is manufactured and used under strictly controlled conditions over the entire lifecycle. Exposure is limited to occasional sampling tasks for quality control, as well as to charging and discharging processes. Here, as well as during the handling for quality control analysis in the laboratory, risk management measures and stringent personal protective equipment are continuously applied. Transport, storage tanks, reactors, processing equipment, and feeds operate in fully closed systems.

  

Procedural and control technologies are used to minimize residual emissions/exposure as well as qualitative risk considerations:

Operational and technical conditions and measures affecting and controlling workers exposure, such as local exhaust ventilation as well as personal protective equipment, such as goggles, chemically resistant gloves, and respiratory protection where potential exposure may occur as reported in the CSR are followed (see chapters 9 & 10).

 

Based on the described process conditions, testing of 3-Aminopropanol (3-AP, CAS 156-87-6) in a repeated dose oral toxicity study (OECD 407/408) was not performed since the criteria of exposure-based adaptation of information requirements are met.

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. The substance is not considered to be classified for repeated dose toxicity according to Regulation (EC) No 1272/2008 (CLP), as amended for the tenth time in Regulation (EU) No 2017/776.