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Description of key information

The calculated oral and dermal LD50 for tris[(2-hydroxyethyl)ammonium] citrate are > 2000 mg/kg bw, based on the read-across with monoethanolamine and citric acid. 

Key value for chemical safety assessment

Acute toxicity: via oral route

Endpoint conclusion
Dose descriptor:
LD50
Value:
2 000 mg/kg bw

Acute toxicity: via dermal route

Endpoint conclusion
Dose descriptor:
LD50
Value:
2 000 mg/kg bw

Additional information

No substance-specific data on the acute toxicity of tris[(2 -hydroxyethyl)ammonium] citrate are available. However, according to Article 13 of the REACH legislation, in case no appropriate animal studies are available for assessment, information should be generated whenever possible by means other than vertebrate animal tests, i. e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across.

Tris[(2 -hydroxyethyl)ammonium] citrate is an acidic salt of monoethanolamine and citric acid and is expected to dissociate into the respective monoethanolammonium cation and dihydrogen citrate anion (which may subsequently undergo (partial) dissociation to monohydrogen citrate and citrate anions) upon uptake by the body. Therefore it is considered to be acceptable to derive lacking information on toxicological properties of tris[(2 -hydroxyethyl)ammonium] citrate by read-across from its starting materials. Citric acid is an important intermediate of the Krebs cycle (also known as citric acid cycle), and therefore occurs naturally as a metabolite in virtually all living organisms. It is also used as a natural food preservative and a food additive. The average daily intake (ADI) is not limited according to the evaluation of Joint FAO/WHO Expert Committee on Food Additives (1973), which concluded that the substance is non-hazardous to men. Therefore the toxicological behavior of tris[(2 -hydroxyethyl)ammonium] citrate is expected to be governed primarily by the toxicity of monoethanolamine. Nevertheless available data on citric acid have also been taken into account.

For monoehtanolamine, acute oral, dermal and inhalation toxicity studies are available. In the oral acute toxicity study with rats, performed according to a protocol similar to OECD Guideline 401 (Union Carbide Corporation, 1988a), undiluted monoethanolamine was administered by gavage at dose levels of0.25, 0.50, 1.0, 2.0, 4.0 mL/kg bw (equal to 254, 509, 1018, 2036, 4072 mg/kg bw based on a density of 1.018 g/ml) to groups of male and female rats (5/sex/dose at dose levels up to and including 2.0 mL/kg bw, and 2 males/dose at 4 mL/kg bw dose level). Animals were observed for 14 days and necropsied. All animals in the two highest dose groups died. There were no deaths in 0.25 mL/kg bw group; 1 death (male) in the 0.50 mL/kg bw group and 3 deaths (1 male, 2 females) in 1.0 mL/kg bw group. All deaths occurred relatively rapidly after dosing (within 2 days), except for one male rat that died after 12 days after a dose of 0.5 mL/kg. Both of the male rats receiving monoethanolamine at the maximum peroral dosage of 4.0 mL/kg died after 3 hours. One female in the 2.00 mL/kg group died after 4 hours. The clinical signs included sluggishness, piloerection, slight kyphosis, emaciation, unsteady gait, crust and staining of the fur and unkept appearance. Pathological findings revealed red lungs and distended stomach and intestines filled with red liquid in deceased. Mottled red kidneys were observed in the survivors of 1.0 mL/kg bw group; no remarkable findings were observed in survivors of 0.25 and 0.50 mL/kg bw groups. Based on the results of the study, LD50 in males and females were 1.19 (95% CI = 0.79 - 1.80) ml/kg and 1.07 (95% CI = 0.72 - 1.59) mL/kg. The recalculated LD50 for both sexes was 1089 mg/kg bw.

In the acute dermal toxicity study with rabbits, performed according to a protocol similar to OECD Guideline 402 (Union Carbide Corporation, 1988c), monoethanolamine was administered to groups of 5 male and 5 female rabbits under occlusive dressing for 24 hours at dose levels of 1.0, 2.0 or 4.0 mL/kg bw (equal to 1018, 2036 and 4072 mg/kg bw). Animals were observed for 14 days and then necropsied. No animals died in the lowest dose group. One male and one female animals died in the 2.0 mL/kg bw dose group and 9 animals (5 males, 4 females) died in the 4.0 mL/kg bw group. Erythema, edema, necrosis, scabs formation and ulceration were common skin reactions observed. Salmon-coloured to dark red lungs were observed in deceased animals together with stomach and intestines filled with liquid or gas. The LD50 was 2504 (95% CI = 1822 -3451) mg/kg bw for male and 2881 (95% CI= 1639 -5070) mg/kg day for female rabbits.

In the acute inhalation toxicity study with rats (union Carbide Corporation, 1988b), a group of 5 male and 5 female rats was exposed to the saturated vapour concentration of monoethanolamine, equal to 1.3 mg/L, for 6 hours. There were no deaths or signs of toxicity noted. Pathological findings were uneventful. Based on these findings, LD50 (6 h) for monoethanolamine was > 1.3 mg/L.

For citric acid, acute oral and acute dermal toxicity studies are available. In the acute oral toxicity study with mice, performed by protocol similar to OECD Guideline 401 (Roche, 1981), groups of 5 mice of each sex were given a single gavage dose of 0, 3, 4.2, 6, 8.5 and 13 g/kg bw with observation for mortality for 10 days. Clinical observations were made at 2 h and 24 h after treatment; body weights were only recorded prior to dosing. There were no mortalities in the controls and 3 g/kg bw group. All animals in the 8.5 and 13 g/kg bw groups died. The mortality in 4.2 and 6 g/kg bw group were 2 and 7 out of 10 animals, respectively. Based on the results of the study, LD50 of 5.4 g/kg bw was determined for citric acid.

In the acute dermal toxicity study with rats, performed according to OECD guideline 402 and under GLP (Safepharm, 2006), a group of 5 male and 5 female rats received a single application of 2000 mg/kg bw test substance for 24 hours under semi-occlusive conditions. The animals were observed for deaths and overt signs of toxicity 1/2, 1, 2 and 4 hours after dosing. Individual bopdy weights were recorded prior to application of the test material on day 0 and on days 7 and 14. There were no deaths or signs of systemic toxicity. No abnormalities were recorded at necropsy. Based on the results of the study, the LD50 was established to exceed 2000 mg/kg bw.

As citric acid is obviously practically non-toxic upon oral and dermal exposure, the acute toxicity data for monoethanolamine will be used as a starting point for the read-across. As the toxicity of tris[(2 -hydroxyethyl)ammonium] citrate is expected to be caused by the toxicity of monoethanolamine released upon dissociation, a correction for molecular weight is warranted. A three times molar equivalent of monoethanolamine is produced upon the intake of tris[(2-hydroxyethyl)ammonium] citrate. Using the oral LD50 value of 1089 mg/kg bw and applying a correction factor of (375.37/3)/61.08, corresponding to the ratio of the molecular weight of tris[(2-hydroxyethyl)ammonium] citrate and three molar equivalents of monoethanolamine,an LD50 of 2231 mg/kg bw is calculated for tris[(2-hydroxyethyl)ammonium] citrate. For acute dermal toxicity, using the lower value of 2504 mg/kg bw for male rabbits, a value of 2504 × (375.37/3)/61.08 = 5129 mg/kg bw is obtained. Both values exceed the maximum limit doses recommended by OECD Guidelines and are far above the classification limits of 2000 mg/kg bw for acute toxicity. Thus classification of tris[(2-hydroxyethyl)ammonium] citrate for acute toxicity is not warranted in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008 and EU Directive 67/548/EEC. As the calculated values are excessively high and may exceed the maximal tolerated physiological dose, the OECD guideline limit values of 2000 mg/kg bw will be taken forward for risk assessment.

 

No LC50 value can be derived for acute inhalation toxicity based on the available data. However, in accordance with Annex VIII of REACH, this information is not required, as data on two routes of exposure are available.

Justification for classification or non-classification

Based on the calculated LD50 values for acute oral and dermal toxicity for tris[(2 -hydroxyethyl)ammonium] citrate, classification is not warranted in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008 and EU Directive 67/548/EEC.