Bis(C12-C13)alkyl-2-hydroxybutandioate

Endpoint summary

Description of key information

Short description of key information on bioaccumulation potential result: 
Bis (C12 -C13)alkyl-2 -hydroxybutanedioate will be hydrolysed by esterases and cleaved into isotridecyl alcohol and adipic acid. Isotridecyl alcohol will be oxidized to the corresponding aldhyde and subsequently to the carboxylic acid.
Carboxylic acid will undergo oxidation (beta and at other positions). beta-Oxidation products will  be subject to fatty acid degradation and ultimately enter the citric acid cycle. Due to its bulky branched structure, other metabolic transformations as alcohol oxidation and fatty acid beta-oxidation are expected play an important role resulting in hydroxy acids, dialcohols and dicarboxylic acids. Conjugation of various metabolites is very likely in order to solubilise the highly hydrophobic sceletal structure.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

Bis (C12 -C13)alkyl-2 -hydroxybutanedioate

The substance is a diester of a dicarboxylic acid consisting of 2 -hydroxybutanoic acid (C4 carbon frame) and isododecyl alcohols and isotridecyl alcohols with a branched carbon chain (n = 13 and n = 12). Accordingly, the substance is expected to show the typical characteristics of an ester. In vivo, esters are hydrolysed by ubiquitous hydrolases to alcohol and carboxylic acid. In case of diesters, this operation will proceed in two steps resulting first in a monoester. Isododecyl alcohol and Isotridecyl alcohol and 2 -hydroxybutanoic acid, formed by hydrolysis of the substance, will then follow metabolic pathways typical for alcohols and carboxylic acids.

 

In principle given the background information on aliphatic saturated long chain esters, alcohols and acids, the following properties and metabolic pathways are expected for Bis (C12 -C13)alkyl-2 -hydroxybutanedioate.

Absorption:

Bis (C12 -C13)alkyl-2 -hydroxybutanedioate and eventually isododecyl alcohol and isotridecyl alcohol and 2 -hydroxybutanoic acid are rapidly absorbed from the gastro-intestinal tract, whereas dermal absorption is expected to be slow.

Biotransformation: The substance is expected to hydrolyse rapidly in vivo to isododecyl alcohol and isotridecyl alcohol and 2 -hydroxybutanoic acid. The alcohol will be oxidised by alcohol dehydrogenase and aldehyde dehydrogenase to the corresponding aldehyde and carboxylic acid. Isododecyl alcohol and Isotridecyl alcohol are not expected to be good substrates for ADH and AlDH, due to their branched bulky structure. ß-Oxidation of the carboxylic acid may be hindered by methy/alkyl substituents at uneven positions, forcing oxidation at other positions. and preventing further degradation in the citrate cycle. Therefore, significant chain hydroxylation and conjugation reactions of the alcohol and of other metabolic oxidation products are expected to account for the majority of the biotransformation.

Excretion of polar metabolites and conjugates may occur via urine and bile and is estimated to be substantial. Entero-hepatic circulation of metabolites excreted via bile is likely to occur (Eisenbrand 2002).

Metabolite malic acid

Male albino Wistar rats were given radioactive marked Malic acid in an aqueous solution by gavage or by intraperitoneal injection. Most of the radioactiivity was excreted as carbon dioxide, after 24 hours 91.6 % and 83.4 % of orally and intraperitoneally administered substance. After oral administration 3.1 % were found in the urine and 0.6 % in the faeces, after intraperitoneal administration 8.8 % in the urine and 1.4 % in the faeces (International Journal of Toxicology, 1991).

Malic acid is an intermediate in the tricarboxylic acid (Krebs) cycle. It is oxidised to oxaloacetic acid and plays an essential role in carbohydrate metabolism (International Journal of Toxicology, 1991).

Metabolites Alcohols, C12 -C13, branched and linear

The initial step in the mammalian metabolism of primary alcohols is the oxidation to the corresponding carboxylic acid, with the corresponding aldehyde being a transient intermediate. These carboxylic acids are susceptible to further degradation via acyl-CoA intermediates by the mitochondrial ß-oxidation process. This mechanism removes C2 units in a stepwise process and linear acids are more efficient in this process than the corresponding branched acids. In the case of unsaturated carboxylic acids, cleavage of C2-units continues until a double bond is reached. Since double bonds in unsaturated fatty acids are in the cis-configuration, whereas the unsaturated acyl-CoA intermediates in the ß-oxidation cycle are trans, an auxiliary enzyme, enoyl-CoA isomerase catalyses the shift from cis to trans. Thereafter, ß-oxidation continues as with saturated carboxylic acids (WHO, 1999). An alternative metabolic pathway for aliphatic acids exists through microsomal degradation via Omega-or Omega–1 oxidation followed by β-oxidation. This mechanism provides an efficient stepwise chain-shortening pathway for branched aliphatic acids (Verhoeven, et al., 1998). The acids formed from the longer chained aliphatic alcohols can also enter the lipid biosynthesis and may be incorporated in phospholipids and neutral lipids (Bandi et al, 1971and Mukherjee et al. 1980). A small fraction of the aliphatic alcohols may be eliminated unchanged or as the glucuronide conjugate (Kamil et al., 1953). Similar to the dermal absorption potential, it is expected that orally administered aliphatic alcohols also show a chain-length dependant potential for gastro-intestinal absorption, with shorter chain aliphatic alcohols having a higher absorption potential than longer chain alcohols. With regards to the blood-brain barrier a chain-length dependant absorption potential exists with the lower aliphatic alcohols and acids more readily being taken up than aliphatic alcohols/acids of longer chain-length (Gelman, 1975). Taking into account the efficient biotransformation of the alcohols and the physico-chemical properties of the corresponding carboxylic acids the potential for elimination into breast milk is considered to be low. The long chain aliphatic carboxylic acids are efficiently eliminated and aliphatic alcohols are therefore not expected to have a tissue retention or bioaccumulation potential (Bevan, 2001). Longer chained aliphatic alcohols within this category may enter common lipid biosynthesis pathways and will be indistinguishable from the lipids derived from other sources (including dietary glycerides) (Kabir, 1993; 1995a,b). A comparison of the linear and branched aliphatic alcohols shows a high degree of similarity in biotransformation. For both sub-categories the first step of the biotransformation consists of an oxidation of the alcohol to the corresponding carboxylic acids, followed by a stepwise elimination of C2 units in the mitochondrial β-oxidation process. The metabolic breakdown for both the linear and mono-branched alcohols is highly efficient and involves processes for both sub-groups of alcohols. The presence of a side chain does not terminate the β-oxidation process, however in some cases a single Carbon unit is removed before the C2 elimination can proceed. In summary, long chained alcohols are generally highly efficiently metabolised and there is limited potential for retention or bioaccumulation for the parent alcohols and their biotransformation products.

Discussion on bioaccumulation potential result:

For bis (C12 -C13)alkyl-2 -hydroxybutanedioate no data on toxicokinetics could be identified. Based on the structure of bis (C12 -C13)alkyl)-2 -hydroxybutanedioate, basic information on properties of aliphatic carboxylic esters is used to characterise the substance (Semino,

1989 a, b) (Eisenbrand 2002).

Bis (C12 -C13)alkyl-2 -hydroxybutanedioate

The substance is a diester of a dicarboxylic acid consisting of 2 -hydroxybutanoic acid (C4 carbon frame) and isododecyl alcohols and isotridecyl alcohols with a branched carbon chain (n = 13 and n = 12). Accordingly, the substance is expected to show the typical characteristics of an ester. In vivo, esters are hydrolysed by ubiquitous hydrolases to alcohol and carboxylic acid. In case of diesters, this operation will proceed in two steps resulting first in a monoester. Isododecyl alcohol and Isotridecyl alcohol and 2 -hydroxybutanoic acid, formed by hydrolysis of the substance, will then follow metabolic pathways typical for alcohols and carboxylic acids.

 

In principle given the background information on aliphatic saturated long chain esters, alcohols and acids, the following properties and metabolic pathways are expected for bis (C12 -C13)alkyl-2 -hydroxybutanedioate.

Absorption:

Bis (C12 -C13)alkyl-2 -hydroxybutanedioate amd eventually isododecyl alcohol and isotridecyl alcohol and 2 -hydroxybutanoic acid are rapidly absorbed from the gastro-intestinal tract, whereas dermal absorption is expected to be slow.

Biotransformation: The substance is expected to hydrolyse rapidly in vivo to isododecyl alcohol and isotridecyl alcohol and 2 -hydroxybutanoic acid. The alcohol will be oxidised by alcohol dehydrogenase and aldehyde dehydrogenase to the corresponding aldehyde and carboxylic acid. Isododecyl alcohol and Isotridecyl alcohol are not expected to be good substrates for ADH and AlDH, due to their branched bulky structure. ß-Oxidation of the carboxylic acid may be hindered by methy/alkyl substituents at uneven positions, forcing oxidation at other positions. and preventing further degradation in the citrate cycle. Therefore, significant chain hydroxylation and conjugation reactions of the alcohol and of other metabolic oxidation products are expected to account for the majority of the biotransformation.

Excretion of polar metabolites and conjugates may occur via urine and bile and is estimated to be substantial. Entero-hepatic circulation of metabolites excreted via bile is likely to occur (Eisenbrand 2002).

Metabolite malic acid

Male albino Wistar rats were given radioactive marked Malic acid in an aqueous solution by gavage or by intraperitoneal injection. Most of the radioactiivity was excreted as carbon dioxide, after 24 hours 91.6 % and 83.4 % of orally and intraperitoneally administered substance. After oral administration 3.1 % were found in the urine and 0.6 % in the faeces, after intraperitoneal administration 8.8 % in the urine and 1.4 % in the faeces (International Journal of Toxicology, 1991).

Malic acid is an intermediate in the tricarboxylic acid (Krebs) cycle. It is oxidised to oxaloacetic acid and plays an essential role in carbohydrate metabolism (International Journal of Toxicology, 1991).

Metabolites Alcohols, C12 -C13, branched and linear

The initial step in the mammalian metabolism of primary alcohols is the oxidation to the corresponding carboxylic acid, with the corresponding aldehyde being a transient intermediate. These carboxylic acids are susceptible to further degradation via acyl-CoA intermediates by the mitochondrial ß-oxidation process. This mechanism removes C2 units in a stepwise process and linear acids are more efficient in this process than the corresponding branched acids. In the case of unsaturated carboxylic acids, cleavage of C2-units continues until a double bond is reached. Since double bonds in unsaturated fatty acids are in the cis-configuration, whereas the unsaturated acyl-CoA intermediates in the ß-oxidation cycle are trans, an auxiliary enzyme, enoyl-CoA isomerase catalyses the shift from cis to trans. Thereafter, ß-oxidation continues as with saturated carboxylic acids (WHO, 1999). An alternative metabolic pathway for aliphatic acids exists through microsomal degradation via Omega-or Omega–1 oxidation followed by β-oxidation. This mechanism provides an efficient stepwise chain-shortening pathway for branched aliphatic acids (Verhoeven, et al., 1998). The acids formed from the longer chained aliphatic alcohols can also enter the lipid biosynthesis and may be incorporated in phospholipids and neutral lipids (Bandi et al, 1971aandb and Mukherjee et al. 1980). A small fraction of the aliphatic alcohols may be eliminated unchanged or as the glucuronide conjugate (Kamil et al., 1953). Similar to the dermal absorption potential, it is expected that orally administered aliphatic alcohols also show a chain-length dependant potential for gastro-intestinal absorption, with shorter chain aliphatic alcohols having a higher absorption potential than longer chain alcohols. With regards to the blood-brain barrier a chain-length dependant absorption potential exists with the lower aliphatic alcohols and acids more readily being taken up than aliphatic alcohols/acids of longer chain-length (Gelman, 1975). Taking into account the efficient biotransformation of the alcohols and the physico-chemical properties of the corresponding carboxylic acids the potential for elimination into breast milk is considered to be low. The long chain aliphatic carboxylic acids are efficiently eliminated and aliphatic alcohols are therefore not expected to have a tissue retention or bioaccumulation potential (Bevan, 2001). Longer chained aliphatic alcohols within this category may enter common lipid biosynthesis pathways and will be indistinguishable from the lipids derived from other sources (including dietary glycerides) (Kabir, 1993; 1995a,b). A comparison of the linear and branched aliphatic alcohols shows a high degree of similarity in biotransformation. For both sub-categories the first step of the biotransformation consists of an oxidation of the alcohol to the corresponding carboxylic acids, followed by a stepwise elimination of C2 units in the mitochondrial β-oxidation process. The metabolic breakdown for both the linear and mono-branched alcohols is highly efficient and involves processes for both sub-groups of alcohols. The presence of a side chain does not terminate the β-oxidation process, however in some cases a single Carbon unit is removed before the C2 elimination can proceed. In summary, long chained alcohols are generally highly efficiently metabolised and there is limited potential for retention or bioaccumulation for the parent alcohols and their biotransformation products.

 

Di-2-ethylhexyl adipate (DEHA)

Absorption, Distribution, Excretion

An ADME study was performed in mice, rats and monkeys (CMA, 1984). The data showed that after oral administration DEHA or its metabolites is readily absorbed, and distributed to various tissues (with highest levels recovered in blood and liver). From these data absorption rates of 67-98% were noted. The highest absorption rates were noted for the mice 98 -100%, in comparison with rats (75%) and monkeys (49-67%).

The excretion rates were also extensive (90-100%) and rapid. After 24 hours rats and monkeys showed lower elimination in urine (60-74%) and higher elimination in feces (app. 20 %).

Metabolism

Urine of mice, rats, and monkeys contained 2-ethylhexanoic acid (EHA), its glucuronic acid conjugate, a hydroxy acid (5-hydroxy-2-ethylhexanoic acid,5-OH EHA), and the diacid (2-ethyl-1,6-hexanedioic acid, DiEHA). In monkeys, glucuronides of the monoester, monoethylhexyladipate (MEHA), and the alcohol, ethylhexanol (EH) were also tentatively identified. In a human volunteers study, similar metabolites were found in urine and MEHA was found in fecal samples (Loftus, 1993).

The GI-tracts contained appreciable amounts of Diester (DEHA), monoester, and alcohol (EH). Hydrolysis to the alcohol through the monoester appears, therefore, to be the principal metabolic process.

Although the site of DEHA metabolism was not definitively determined, the data suggest that hydrolysis to the mono-ester occurs primarily in the GI tract. The absorbed 14C appears to be primarily the alcohol; the diester and monoester were detected in the liver only in small quantities.

Resumé

Overall the data presented in the ADME studies indicate that orally administered 14C-DEHA was rapidly hydrolyzed in the GI tract before absorption and distribution to tissues. Hydrolysis of the monoester, MEHA, also appears to occur readily, leading to the low recovery of this metabolite in the hepatic tissue and urine. Low doses of' DEHA and/or its metabolites were absorbed rapidly and distributed to major organs and tissues. Absorption, however, was incomplete following a high dose of 14C-DEHA. The absorbed radioactivity is metabolized further in the liver before elimination in urine, expired air, and feces. Dose-dependent changes in absorption, tissue uptake, metabolism, and elimination were demonstrated in mice. Sex differences were also apparent in the hepatic uptake and metabolism of the absorbed radioactivity. The data indicate little, if any, prolonged retention of DEHA or its metabolites in blood and tissue following oral administration to mice, rats, and monkeys.

 

Justification of use of di-2 -Ethylhexyl adipate (DEHA) as supporting substance for Bis(C12 -C13)alkyl-2 -hydroxybutandioate for read across

The toxicokinetics study on DEHA confirms the concept of bis(C12 -C13)alkyl-2 -hydroxybutandioate

metabolism developed based on background information. Ester hydrolysis proceeds in two steps. Hydroxy- and di- and monocarboxylic acids are formed. There is conjugation of distinct oxidised metabolites and expired CO2 is quite low.

Both substances have the same basic structure (diester of an acid, difference in the chain length C-6 versus C-4). There are differences in the chain length of the alcohol part of the esters (C-12, 13 versus C-8). This variation is not considered to cause substantial changes in the biological effects of the esters. Metabolism can be expected to be basically the same, likely with only variations in the amount and distribution of individual metabolites. Due to its structural and chemical closeness, both substances can be expected to behave similar if administered to experimental organisms and animals but it is additionally something like a "worst case" because of the "2 -ethylhexyl"-group which is a structure of toxicological concern (e.g. for the endpoint Reproduction Toxicity). Thus it is justified to use di-ethylhexyl adipate as supporting substance for Bis(C12 -C13)alkyl-2 -hydroxybutandioate.

 

The same argumentation is valid if ditridecyl adipate (DTDA) are used as supporting substance for cross reading.

Justification of use of Malic acid and Alcohol, C12 -C13 , branched and linear, as supporting substance for Bis(C12 -C13)alkyl-2 -hydroxybutandioate

It is expected that in the first step of the metabolism of Bis(C12 -C13)alkyl-2 -hydroxybutandioate the metabolites Malic acid and Alcohol, C12 -C13, branched and linear occur because of the hydrolysis of the ester. Therefore also some toxicological data of these substances were cited and discussed as supporting data.