2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

absorption

Principles of method if other than guideline:

The mechanism of the intestinal fat absorption has been studied with 14C labeled fat in rats with the intestinal lymph duct cannulated.

GLP compliance:

no

Radiolabelling:

yes

Remarks:

14C labeled fat

Species:

rat

Strain:

not specified

Sex:

not specified

Route of administration:

oral: gavage

Duration and frequency of treatment / exposure:

single oral exposure(at least 18 hours after surgery)

Remarks:

Doses / Concentrations:A) 0.5 mL corn oil + 2.5 mg active palmitic acid-1-14CB) 0.5 mL corn oil transesterified with 2.5 mg active palmitic acid-1-14CC) 0.5 mL hydrolysed corn oil + 2.5 mg active palmitic acid-1-14C

No. of animals per sex per dose / concentration:

5-6

Control animals:

no

Details on absorption:

24 hours after administration of the different fats the mean recovered activities in lymph were as following: A) 0.5 mL corn oil + 2.5 mg active palmitic acid-1-14C: 57.0 %B) 0.5 mL corn oil transesterified with 2.5 mg active palmitic acid-1-14C: 61.7 %C) 0.5 mL hydrolysed corn oil + 2.5 mg active palmitic acid-1-14C: 62.3 % In all three groups of experiments maximum recoveries were found after 24 hours, i.e. 80.9, 85.0 and 87.5 % of the activity given.Free fatty acids administered alone or together with glycerides appear in the lymph in glycerides and phospholipids. No free fatty acids or soaps appear in the lymph. The intestinal wall supplies a quantitatively important part of phospholipids to the blood during fat absorption. The recoveries in the lymph of the fat fed varied widely. Diarrhea occured in some animals especially after feeding hydrolysed corn oil.

Details on distribution in tissues:

Absorbed fat is mainly transported via lymphatic channels to the systemic circulation whether fed as glycerides or as fatty acids.

Details on metabolites:

A complete hydrolysis of the fat in the intestinal lumen might occur in the rat.

The proportions of neutral fat and phospholipids in the lymph were in all three cases about the same. 90% of the fatty acids were present in the neutral fat and the remaining 10 % in phospholipids. The neutral fat consisted chiefly of triglycerides; cholesterol and cholesterol esters representing only a minor part of this fraction. No free fatty acids or soaps appeared in the lymph.

The results indicated that glycerides might be completely hydrolysed in the intestinal lumen of the rat and then resynthesized in the intestinal wall.

Conclusions:

Mean absorption rate of corn oil combined with palmitic acid was between 57 - 62 %.

Reference 2

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented publication meeting basic scientific principles

Objective of study:

metabolism

Principles of method if other than guideline:

The lipolytic activity of human gastric and duodenal juice against medium chain and long chain triglycerides was compared.

GLP compliance:

no

Radiolabelling:

yes

Remarks:

Glyceryl trioctanoate-1-14C

Species:

human

Route of administration:

other: in vitro testing

Enzymatic Lipolysis by Gastric and Duodenal Juice:

All samples of gastric juice showed lipolytic activity against trioctanoin and triolein. Hydrolysis of emulsified trioctanoin was greater than of emulsified triolein. Hydrolysis of unemulsified trioctanoin was less and more variable.

Duodenal juice was more active, even against unemulsified trioctanoin and triolein. Duodenal juice was more active against unemulsified substrate than gastric juice against emulsified substrate.

Table 1: Hydrolysis of trioctanoin and triolein*

	Substrate and form (μmoles)	Hydrolysis (%)
		Trioctanoin	Triolein
Gastric juice	30, unemulsified	21	1
	60, emulsified	33	16
Duodenal juice	30, unemulsified	40	34
	45, emulsified	42	35
	105, emulsified	45	36

*Gastric or duodenal juice (1 mL) was incubated (1 hour, continuous shaking, 37ºC) with 1 mL of buffer and unemulsified substrate or 1 mL of substrate emulsified in 10 mM sodium taurodeoxycholate, pH6.

pH Optimum

In the presence of bile acids, gastric lipolytic activity against trioctanoin had a broad pH optimum, between 4 and 7. The lipolytic activity of duodenal juice had a sharper pH optimum, between 6 and 8. The pH optimum was lower for short chain triglycerides, indicating that pH optimum values for lipases must be defined for a particular substrate.

Chain Length Specificity

Lipolysis rates increased with decreasing chain lengths for pure triglycerides.

Tributyrin was cleaved more rapidly than trihexanoin which in turn was cleaved more rapidly than trioctanoin (ratio of rates, 100:69:53). Because the pH optimum of gastric lipase is lower for short chain triglycerides than for MCT, trihexanoin and tributyrin were cleaved much more rapidly than, for example, trioctanoin at pH5.

Esterification and Fatty Acid Acceptors by Gastric and Duodenal Lipases

Gastric and duodenal lipases did not induce esterification of the fatty acid acceptor, glyceryl 2 -monooleyl ester, by octanoic acid over the pH range of 2 to 6. However, it was esterified by oleic acid in the presence of gastric juice, duodenal juice, or pancreatic fistula juice when bile acids were added. Esterification, calculated by disappearance of titratable fatty acid, was confirmed by TLC which showed the formation of compounds having the mobilities of a monoether monoester and a monoether diester. Control incubations without enzyme showed no loss of oleic acid or appearance of new lipids by TLC. To determine the amount of disubstituted and trisubstituted glyceryl derivatives which were formed, 14C-labeled glyceryl 2 -monooleyl ether was used and the products of the reaction were examined by zonal scanning. The glyceryl 2 -monooleyl ether was not cleaved during the incubation procedure. The amounts of ester bonds formed estimated by titration and by zonal scanning were in good agreement.

Products of Lipolysis and Positional Specificity

The specificity of pancreatic lipase for the 1 -ester bond in LCT has been demonstrated previously by establishing the formation of 2 -monoglycerides and fatty acid as end products of lipolysis. This procedure cannot be used for MCT because medium chain 2 -monoglycerides are either cleaved by pancreatic lipase or rapidly isomerized to the 1 -isomer which is rapidly hydrolyzed or both. Indeed, chromatographic examination of the products of hydrolysis of trioctanoin-14C showed only a small fraction of monoglyceride present.

Table 2: Products of hydrolysis of trioctanoin by gastric juice*

	Radioactivity distribution** (%)				Lipolysis (%)
	Monoglyceride	Diglyceride	Fatty acid	Triglyceride
Buffer (control)	0	0	0	100	0
Gastric juice 1 mL	3	26	26	44	34
	3	28	24	43	33
	4	28	25	43	36
	4	28	25	43	36
Duodenal juice
0.4 mL	4	9	15	72	26
0.5 mL	4	14	20	62	40

*Glyceryl trioctanoate-1-¹⁴C was added to 1 mL of emulsified trioctanoin (60 μmoles) and incubated for 1 hour at 37ºC with buffer (blank) or gastric or duodenal juice. The reaction mixture was extracted and a 50 μL aliquot was analyzed by TLC and zonal scanning. A 3 mL aliquot was titrated to quantify fatty acids liberated.

Discussion:

The work confirmed extensive literature showing that gastric juice contains lipolytic activity, that ingested triglyceride is hydrolyzed in the stomach, even after pancreatic diversion, that lipase may be demonstrated histochemically in gastric mucosa, and that gastric mucosal homogenates have lipolytic activity. Pancreatic lipase has some activity at the pH of gastric content, which is between pH6 and pH3 in normal subjects.

Reference 3

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented publication meeting basic scientific principles.

Objective of study:

absorption

Principles of method if other than guideline:

The absorbability of the fatty acid moiety of the complete, oleate esters of alcohols containing from one to six hydroxyl groups was determined by the fat balance technique in adult rats. Similarly, the absorbability of sucrose octaoleate and sucrose monooleate was determined.

GLP compliance:

no

Radiolabelling:

no

Species:

rat

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS- Source: no data- Age at study initiation: young adult- Weight at study initiation: approx. 200 g- Housing: Individually in cages with raised screen bottoms- Diet (e.g. ad libitum): ad libitum

Route of administration:

oral: feed

Duration and frequency of treatment / exposure:

10 Days, diet ad libitum

Remarks:

Doses / Concentrations:10% and 25 % of dietary fat

Details on absorption:

The fatty acids of the compounds containing less than four ester groups, methyl oleate, ethylene glycol dioleate, glycerol trioleate, and sucrose monooleate, were almost completely absorbed. As the number of ester groups was increased - erythritol and pentaerythritol tetraoleate and xylitol pentaoleate - the absorbability decreased. The fatty acids of sorbitol hexaoleate and sucrose octaoleate were not absorbed. These differences in absorbability are related to the activity and specificity of the lipolytic enzymes in the lumen of the intestinal tract.

The fatty acids of the compounds containing less than four ester groups, methyl oleate, ethylene glycol dioleate, glycerol trioleate, and sucrose monooleate, were almost completely absorbed. As the number of ester groups was increased - erythritol and pentaerythritol tetraoleate and xylitol pentaoleate - the absorbability decreased. The fatty acids of sorbitol hexaoleate and sucrose octaoleate were not absorbed. These differences in absorbability are related to the activity and specificity of the lipolytic enzymes in the lumen of the intestinal tract.

Test fat	Percentage of dietary fat	Absorbability [%]
Methyl Oleate	10	100
	25	96
Ethylen Glycol Oleate	10	100
	25	92
Glycerol Trioleate	100	(100)
Erythritol Tetraoleate	10	-
	25	72
Pentaerythritol Tetraoleate	10	90
	25	64
Xylol Pentaoleate	10	50
	25	24
Sorbitol hexaoleate	10	0
	25	0
Sucrose Octaoleate	5	0
	10	0
	15	0
Sucrose Monooleate	5	100
	10	100
	15	100

Conclusions:

Absorption rates were between 0 an 100 %, depending on the amount of ester groups present in the substance fed. Pentaerythritole tetraoleate had an absorption rate of 90% (10% of dietary fat) and 64% (25% of dietary fat), respectively. Erythritole tetraoleate had an absorption rate of 72% (25% of dietary fat).

Reference 4

Endpoint:

basic toxicokinetics

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Only secondary dataShort review on metabolism from previous publications.

Objective of study:

metabolism

The metabolism of Medium chain triglycerides in the canine is a process whereby lipases from the buccal cavity and pancreas release the fatty acids in the gastrointestinal tract where they are absorbed. Unlike long chain triglycerides (LCT), where long chain fatty acids (LCFA) form micelles and are absorbed via the thoracic lymph duct, MCFA are most often transported directly to the liver through the portal vein and do not necessarily form micelles. Also, MCFA do not re-esterify into MCT across the intestinal mucosa. MCFA are transported into the hepatocytes through a carnitine-independent mechanism, and are metabolized into carbon dioxide, acetate, and ketones through b-oxidation and the citric acid cycle.

Description of key information

Absorption of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate may be the highest after oral exposure, but may be limited at all due to the low water solubility. After enzymatic hydrolysis, the hydrolysis products may be absorbed and well distributed within the body. Alcohol hydrolysis products are incorporated in standard metabolic pathways. 2-Ethylhexanoic acid may be metabolised by omega and omega-1 oxidation or by glucuronidation and following renal excretion.

Additional information

In accordance with Annex VIII, Column 1, Item 8.8 of Regulation (EC) 1907/2006 and with Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance (ECHA, 2008), assessment of the toxicokinetic behaviour of the substance 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate (CAS No. 28510-23-8) was conducted to the extent that can be derived from the relevant available information on physicochemical and toxicological characteristics. There are no studies evaluating the toxicokinetic properties of the substance available. Some information is available for the hydrolysis product ethylhexanoic acid and other substances with similar chemical moieties.

Absorption

Absorption is a function of the potential of a substance to diffuse across biological membranes. The most useful parameters providing information on this potential are the molecular weight, the octanol/water partition coefficient (log Pow) value and the water solubility. The log Pow value provides information on the relative solubility of the substance in water and lipids (ECHA, 2008).

Oral

When assessing the potential of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate to be absorbed in the gastrointestinal (GI) tract, it has to be considered that carboxylic acid esters will undergo enzymatic hydrolysis by ubiquitously expressed GI esterases (Long, 1958; Lehninger, 1970; Mattson and Volpenhein, 1972). The rate of hydrolysis is dependent on the structure of the ester, and may therefore be rapid or rather slow.

Thus, due to hydrolysis predictions on oral absorption based on the physico-chemical characteristics of the intact parent substance alone may no longer apply. Instead, the physico-chemical characteristics of the breakdown products of the ester (neopentylglycol and the carboxylic acid 2-ethylhexanoic acid) may become relevant. The molecular weight of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate (356 g/mol) does favour absorption. In contrast, the low water solubility (0.01 mg/L) and the high log Pow value of the parent compound (7.5) indicate that the absorption may be limited by the inability to dissolve into GI fluids. However, micellar solubilisation by bile salts may enhance absorption, a mechanism which is especially of importance for highly lipophilic substances with log Pow >4 and low water solubility (Aungst and Shen, 1986).

When considering the hydrolysis product neopentylglycol, absorption will occur, as oral administration of neopentylglycol led to urinary excretion of neopentylglycol or its metabolites (Gessner et al., 1960).

The other hydrolysis product 2-ethylhexanoic acid (molecular weight 144.21 g/mol; water solubility approximately 2 g/L; log Pow approximately 2.7) is well absorbed via the gastrointestinal tract. Peak plasma concentrations of 85.1 µg 2-ethylhexanoic acid equivalents per g blood were reached after 18.8 min following oral administration of 100 mg/kg bw of 2-ethylhexanoic acid (EPA, 1986).

Monoester may also be absorbed after micellar solubilisation by bile salts, similar to mono- and diester of fatty acids with glycerol.

Dermal

There are no data available on dermal absorption or on acute dermal toxicity of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate. On the basis of the following considerations, the dermal absorption of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate is considered to be low. Regarding the molecular weight of 356 g/mol and an octanol/water partition coefficient of 7.5 in combination with the low water solubility, a low dermal absorption rate is anticipated. Log Pow values above 6, will slow the rate of transfer between the stratum corneum and the epidermis and therefore absorption across the skin will be limited and uptake into the stratum corneum itself is slow.

The hazard assessment of the hydrolysis products via the dermal route is secondary, as low hydrolysis of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate in the skin is assumed. 

Inhalation

2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate has a very low vapour pressure of 0.000144 Pa at 20 °C (calculated) thus being of low volatility. Therefore, under normal use and handling conditions, inhalation exposure and thus availability for respiratory absorption of the substance in the form of vapours, gases, or mists is not significant.

However, the substance may be available for respiratory absorption in the lung after inhalation of aerosols, if the formulated substance is sprayed. In humans, particles with aerodynamic diameters below 100 μm have the potential to be inhaled. Particles with aerodynamic diameters below 50 μm may reach the thoracic region and those below 15 μm the alveolar region of the respiratory tract (ECHA, 2008).

As discussed above, absorption after oral administration of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate may mainly be driven by enzymatic hydrolysis of the ester bond to the respective metabolites and subsequent absorption of the breakdown products. As the presence of esterases and lipases in the mucus lining fluid of the respiratory tract is expected to be lower in comparison to the gastrointestinal tract, absorption of the hydrolysis products in the respiratory tract is considered to be less effective than in the gastrointestinal tract. Nevertheless, absorption of the parent substance itself cannot be excluded if the 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate reaches the alveolar region. 

Therefore, inhalative absorption of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate is considered to be not higher than through the intestinal epithelium, but still likely to happen.

Distribution and accumulation

Distribution of a compound within the body depends on the physicochemical properties of the substance especially the molecular weight, the lipophilic character and the water solubility. In general, the smaller the molecule, the wider is the distribution. If the molecule is lipophilic, it is likely to distribute into cells and the intracellular concentration may be higher than extracellular concentration particularly in fatty tissues (ECHA, 2008).

As the parent compound 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate will be hydrolysed before absorption as discussed above, the distribution of intact 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate is not relevant but rather the distribution of the breakdown products of hydrolysis. The absorbed products of hydrolysis, neopentylglycol and 2-ethylhexanoic acid can be both distributed within the body.

Data from studies in animals indicate neopentylglycol is rapidly absorbed in the intestine and the stomach and excreted mainly in the urine (62% within 24 h).

In summary, the available information on 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate indicate that no significant bioaccumulation of the parent substance in adipose tissue is expected.

Metabolism

Metabolism of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate occurs initially via enzymatic hydrolysis of the ester resulting in neopentylglycol and/or neopentyglycol monoester.

2-Ethylhexanoic acid metabolism was found to take place via conjugation with glucuronic acid as well as cytochrome P-450-dependent omega and omega-1 oxidation. The major urinary metabolites identified were the glucuronide of 2-ethylhexanoic acid as well as 2-ethyl-1,6-hexanedioic acid 6-hydroxy-2-ethylhexanoic acid and their respective glucuronides. With increasing single dose the fraction of glucuronidated 2-ethylhexanoic acid increased while the percentage of cytochrome P-450 dependent, more highly oxidized metabolites decreased.

When considering the hydrolysis product neopentylglycol, absorption will occur. In a toxicokinetic study from 1960 (Gessner et al.), the metabolism of several glycols, including neopentylglycol, was determined. 1-1.5 g/kg bw of neopentylglycol was administered to 4 rabbits by oral gavage. The 24-h urine was analysed for neopentylglycol or metabolism products of neopentylglycol. No other route of excretion was investigated. 62% (range 53-67%) of the applied dose was found in the 24-h urine as the conjugate of glucuronic acid indicating rapid absorption after oral application. 1.9% of the applied dose was excreted as 3-hydroxy-2,2-dimethylpropionic acid and 0.7% as unchanged neopentyl glycol.

In conclusion, rapid absorption of the test substance can be expected followed by conjugation with glucuronic acid and excretion via urine. 

Excretion

Neopentylglycol is well absorbed via the oral route. 62% (range 53-67%) of the applied dose was found in the 24-h urine as the conjugate of glucuronic acid, 1.9% of the applied dose was excreted as 3-hydroxy-2,2-dimethylpropionic acid and 0.7% as unchanged neopentylglycol. No other rote of excretion was investigated.

In conclusion, rapid absorption of the test substance can be expected followed by conjugation with glucuronic acid and excretion via urine.

In all available studies with 2-ethylhexanoic acid, irrespective of the route of administration employed, the radioactivity was predominantly excreted in the urine and faeces within 24 h, with half-lives of elimination ranging from 4.2 to 6.8 h.

A detailed reference list is provided in the technical dossier (see IUCLID, section 13) and within the CSR.