Nonanoic acid, mixed esters with heptanoic acid, pentaerythritol and valeric acid

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Reason / purpose for cross-reference:

reference to same study

Lipids are not only structural building blocks of cells and tissues but at the same time suppliers of C atoms for a number of biosynthetic pathways as well as carriers of essential fatty acids and fat-soluble vitamins. In addition, fatty acids are precursors of prostaglandins and other eicosanoids and therefore have important metabolic functions.

Fatty acids can be divided into three groups, saturated, monounsaturated, and polyunsaturated fatty acids.

Each class of fatty acids has a preferential specific role.

- Saturated fatty acids (medium or long-chain) are more devoted to energy supply, but one should not forget their specific structural role.

- The polyunsaturated fatty acids of the n–3 and n–6 families have very important structural and functional roles and ideally should not be utilized for energy purposes.

 

Table 1:

Role of different classes of fatty acids

Fatty acids	Energy	Structure	Function
Medium-chain saturated fatty acids	+++	0	0
Long-chain fatty acids
Saturated	++	++	(+)
Monounsaturated	++	++	(+)
Polyunsaturated
Linoleic or n-6 family	0	+++	+++
Linolenic or n-3 family	0	+++	+++

 0, +, ++, +++ : Emphasis of contribution, increasing in rank order

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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-1-14C
B) 0.5 mL corn oil transesterified with 2.5 mg active palmitic acid-1-14C
C) 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 3

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

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 4

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

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.

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 5

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Remarks:

Short 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

The hazard assessment is based on the data currently available. New studies with the registered substance and/or other member substances of the polyol esters category will be conducted in the future. The finalised studies will be included in the technical dossier as soon as they become available and the hazard assessment will be re-evaluated accordingly.

For further details, please refer to the category concept document attached to the category object (linked under IUCLID section 0.2) showing an overview of the strategy for all substances within the polyol esters category.

 

Oral absorption

Based on available data, absorption after oral ingestion is predicted to be limited. Due to the rather high number of ester bonds, only slow hydrolysis in the gastrointestinal tract is expected to occur, resulting in hydrolysis products that may be readily absorbed.

Dermal absorption

The low water solubility, the high molecular weight, the high log Pow value and the lack of potential for skin irritation / corrosion indicate that dermal uptake in humans is likely to be low.

Inhalative absorption

A systemic bioavailability in humans after inhalation exposure cannot be excluded, e.g. after inhalation of aerosols with aerodynamic diameters below 15 μm. The absorption rate is not expected to be higher than that following oral exposure.

Distribution and accumulation

The available information indicates that the intact parent compound is not assumed to distribute throughout the body due to limited absorption. In contrast, wide distribution within the body is expected for the hydrolysis products pentaerythritol, dipentaerythritol and the fatty acids. However, no significant bioaccumulation of both the parent substance and its anticipated hydrolysis products in adipose tissue is expected.

Metabolism

Esters of fatty acids are hydrolysed to the corresponding alcohol and fatty acids by ubiquitously expressed esterases. It is assumed, however, that the hydrolysis rate is low as a result of steric hindrance due to the number of ester bonds and the complexity of the parent molecule. If hydrolysis occurs, a major metabolic pathway for linear and simple branched fatty acids is the β-oxidation for energy generation. In contrast, pentaerythritol and dipentaerythritol are absorbed rapidly and mainly excreted unchanged without metabolic transformation.

Excretion

A low absorption rate is expected via the gastrointestinal tract. Thus the biggest part of the ingested substance is assumed to be excreted in the faeces. Following the potential hydrolysis of the parent molecule, the fatty acids are not expected to be excreted to a significant degree via the urine or faeces but excreted via exhaled air as CO2. Pentaerythritol and dipentaerythritol are not metabolised but excreted mainly unchanged via urine.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

The hazard assessment is based on the data currently available. New studies with the registered substance and/or other member substances of the polyol esters category will be conducted in the future. The finalised studies will be included in the technical dossier as soon as they become available and the hazard assessment will be re-evaluated accordingly.

For further details, please refer to the category concept document attached to the category object (linked under IUCLID section 0.2) showing an overview of the strategy for all substances within the polyol esters category.

  

Basic toxicokinetics

There are no studies available in which the toxicokinetic behaviour of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids has been investigated. Therefore, in accordance with Annex VIII, Column 1, Item 8.8 of Regulation (EC) No. 1907/2006 and with the Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance (ECHA, 2017), an assessment of the toxicokinetic behavior was conducted based on relevant available information. This comprises a qualitative assessment of the available substance-specific data on physico-chemical and toxicological properties and taking into account further available information on source substances from which data was used for read-across to cover data gaps.

 

Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids is a UVCB substance as it is characterised by several highly variable parameters. It contains esters of pentaerythritol (PE) and dipentaerythritol (DiPE) with a variety of fatty acids with unknown and/or variable composition. Alkyl chain lengths found in the substance include C5, C7 and C9. Both principal components – the poly-alcohol (PE and DiPE) as well as the fatty acids – are present in varying amounts. The target substance is a liquid with a water solubility of < 2.01 mg/L (Nyco, 2017). The molecular weight of its various constituents ranges between 472.61 - 1095.61 g/mol, the log Pow was estimated to be > 6.5 (QSAR, Vega version 1.1.3 - three models: Meylan/Kowwin version 1.1.4, MLogP version 1.0.0, ALogP version 1.0.0) and the calculated vapour pressure is < 0.0001 Pa at 20 °C (QSAR, ARChem SPARC. version 4.6).

 

General considerations on absorption

Absorption is a function of the potential for 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, 2017).

 

Oral absorption

The smaller the molecule, the more easily it will be taken up. In general, molecular weights below 500 g/mol are favorable for oral absorption (ECHA, 2017). As the molecular weight of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids ranges between 472.61 and 1095.61 g/mol, absorption in the gastrointestinal tract is expected only for the low molecular weight constituents resulting from the C5 fatty acid moiety. C5 fatty acids form about 20 – 40% of all fatty acids contained in the UVCB substance, and therefore it can be expected that the major part of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids will not be readily absorbed after oral exposure. Absorption after oral administration is also rather unexpected when the “Lipinski Rule of Five” (Lipinski et al. (2001), refined by Ghose et al. (1999)) is applied. The log Pow of > 6.5 suggests that the absorption may be limited by the inability to dissolve into the gastrointestinal fluids but be enhanced by micellar solubilisation, as this mechanism is of importance for highly lipophilic substances (with a log Pow > 4), who are poorly soluble in water (1 mg/L or less). However, for large molecules the gastrointestinal absorption is not likely to occur.

 

The potential of a substance to be absorbed from the gastrointestinal tract (GIT) may be influenced by chemical changes taking place in gastrointestinal fluids, for instance due to metabolism by gastrointestinal flora or by enzymes released into the gastrointestinal tract or by hydrolysis. These changes will alter the physico-chemical characteristics of the substance and hence predictions based upon the physico-chemical characteristics of the parent substance may in some cases no longer apply (ECHA, 2017). After oral ingestion, fatty acid esters with glycerol (glycerides) are rapidly hydrolised by ubiquitously expressed esterases and almost completely absorbed (Mattsson and Volpenhein, 1972a). On the contrary, a lower rate of enzymatic hydrolysis in the GIT was demonstrated for compounds with more than 3 ester groups (Mattson and Volpenhein, 1972a,b). The in vitro hydrolysis rate of a pentaerythritol ester was about 2000 times slower in comparison to glycerol esters (Mattson and Volpenhein, 1972a,b). Moreover, in vivo studies in rats demonstrated the incomplete absorption of the compounds containing more than three ester groups. This decrease became more pronounced as the number of ester groups increased, probably the results of different rates of hydrolysis in the intestinal lumen (Mattson and Volpenhein, 1972c). Based on these considerations, polyol esters are capable of being enzymatically hydrolysed to generate alcohol and the corresponding fatty acids. PE and DiPE esters may show different rates of enzymatic hydrolysis depending on the number of ester bonds and the alcohol involved. Nevertheless, the metabolic fate of the substances is considered to be the same, as all polyol ester substances will be hydrolysed over a given period of time.

 

Once hydrolysis took place, the resulting hydrolysis products, fatty acids and the poly- alcohol PE or DiPE, may be absorbed. The highly lipophilic fatty acids will be absorbed by micellar solubilisation (Ramirez et al., 2001). A study by Mattson and Nolen (1972) determined the absorbability of the fatty acid moiety of the complete oleate esters of alcohols containing one to six hydroxyl groups. The fatty acids of the compounds containing less than four ester groups were almost completely absorbed. As the number of ester groups was increased (erythritol and pentaerythritol tetraoleate and xylitol pentaoleate) the absorbability of the compounds decreased but was still present. Polyols (e.g. PE and DiPE) are - due to their physico-chemical properties (low molecular weight, low log Pow, and high solubility in water) - easily absorbed and can either remain unchanged or may be further metabolised or conjugated (e.g. glucuronides, sulfates, etc.) into polar products that are excreted via urine (DiCarlo et al., 1965). The PE, being a highly water-soluble substance (25 g/L; OECD SIDS, 1998), will readily dissolve into the gastrointestinal fluids. 10 mg/kg 14C-labled PE orally administered to mice was absorbed rapidly. Almost half of the administered dose left the gastrointestinal tract within 15 minutes (DiCarlo et al., 1965). A similar behaviour is expected for DiPE since its water solubility is much higher (3000 mg/L; OECD SIDS, 1995).

 

The available data on acute and repeated dose oral toxicity support the conclusion of only limited oral absorption. Acute oral toxicity investigations have been performed with the source substances Pentaerythritol ester of pentanoic acids and isononanoic acid (CAS No. 146289-36-3), Pentaerythritol tetraesters of n-decanoic, n-heptanoic, n-octanoic and n-valeric acids (CAS No. 68424-31-7) and Fatty acids, C5-9 tetraesters with pentaerythritol (CAS No. 67762-53-2). A single administration of 2000 mg/kg bw test material to male and female rats did not induce any mortality or any signs of systemic toxicity (Henkel, 1991; ICI, 1991a; Huntington, 1999a; Mobil, 1984). Moreover, in subacute and subchronic repeated dose toxicity studies performed with the source substances Pentaerythritol tetraesters of n-decanoic, n-heptanoic, n-octanoic and n-valeric acids (CAS No. 68424-31-7), Fatty acids, C8-10 mixed esters with dipenaterythritol, isooctanoic acid, pentaerythritol and tripentaerythritol (CAS No. 189200-42-8) and Pentaerythritol ester of pentanoic acids and isononanoic acid (CAS No. 146289-36-3), oral exposure of male and female rats either in the diet or by gavage did not yield any adverse effects and hence resulted in NOAEL values of ≥ 1000 mg/kg bw/day, corresponding to the highest doses tested (Zeneca, 1993; Exxon, 1995d; ToxLab, 1998). However, it must be noted that the lack of systemic toxicity observed in the studies can also be attributed to a low degree of toxicity. The lack of systemic toxicity is therefore only an indicator rather than a proof of no or a low absorption potential.

 

Based on the physico-chemical properties of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids as well as on available literature and data obtained with structural analogue substances, absorption after oral ingestion is predicted to be limited. Due to the rather high number of ester bonds, only slow hydrolysis in the GIT is expected to occur, resulting in hydrolysis products that may be readily absorbed. If absorption of the intact parent compound or the respective metabolites occurs, it is predicted to cause no or only a low order of systemic toxicity. This assumption is also supported by available data on oral toxicity.

 

Dermal absorption

Similar to oral absorption, dermal absorption is favored for small molecules. In general, a molecular weight below 100 g/mol favors dermal absorption, while a molecular weight above 500 g/mol may be considered too large (ECHA, 2017). As the molecular weight of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids ranges between 472.61 and 1095.61 g/mol, a dermal absorption even of the low molecular weight constituents of the substance is not likely.

 

If a substance is a skin irritant or corrosive, damage to the skin surface may enhance penetration through the skin. Since Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids was not tested for skin irritation or corrosion, read-across from a variety of structural analogue substances was applied. The structurally related source substances chosen cover carbon-chain distributions of C5 to C10 with linear as well as branched alkyl moieties. None of the source substances was identified as skin irritant or corrosive to skin (ICI, 1991b; Huntington, 1999c; Exxon, 1995a). Therefore, also the target substance is not considered to be irritating or corrosive to skin in humans and an enhanced penetration of the substance due to local skin damage can be excluded.

 

For substances with a log Pow above 4, the rate of dermal penetration is limited by the rate of transfer between the stratum corneum and the epidermis, but uptake into the stratum corneum will be high. For substances with a log Pow above 6, the rate of transfer between the stratum corneum and the epidermis will be slow and will limit absorption across the skin, and the uptake into the stratum corneum itself is also slow. The substance must be sufficiently soluble in water to partition from the stratum corneum into the epidermis (ECHA, 2017). As the water solubility of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids is less than 2.01 mg/L and the log Pow is estimated to be > 6.5, dermal uptake is likely to be very low. The negligible potential for dermal absorption is also supported by QSAR calculations of the dermal absorption rate. Calculations with the Episuite 4.1, DERMWIN 2.02 tool yielded dermal penetration rates of 2.87E-06 (low) and 5.21E-15 mg/cm2/event (very low) for the low molecular weight constituent and the high molecular constituent, respectively.

 

Moreover, the available data on dermal toxicity of the structurally related substance Fatty acids, C5-9 tetraesters with pentaerythritol (CAS No. 67762-53-2) are also considered for the assessment of dermal absorption. In a subchronic (90-day) repeated dose toxicity study, only slight erythema and flaking of the skin were observed in the treated groups. Microscopic examination of the skin indicated trace to slight epidermal hyperplasia and chronic inflammation of the superficial dermis. No signs of systemic toxicity were observed up to and including the highest dose level of 2000 mg/kg bw/day in male and female rats (Mobil, 1988a). The study also investigated the skin penetration potential. The results indicate that the 13-week treatment with the test substance does not increase the skin penetration of the test substance. The skin penetration of untreated rats was less than 2% and the mean value for treated rats was approx. 6%. Furthermore, the same source substance was investigated for its developmental toxicity (Mobil, 1988b). In this study only mild dermal irritation including erythema and flaking of the skin occurred. Since no adverse effects with respect to systemic maternal toxicity were observed, the NOAEL was set at 2000 mg/kg bw/day, corresponding to the highest dose tested.

 

Overall, the calculated low dermal absorption potential, the low water solubility, the high molecular weight (> 100 g/mol), the high log Pow values and the fact that the substance is not irritating or corrosive to skin indicate that dermal uptake of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids in humans can be expected to be limited. This conclusion is further supported by an in vivo dermal penetration investigation of a structural analogue source substance. The experimentally determined skin penetration rate was 2 – 6%.

 

Inhalation absorption

Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids has a low vapour pressure of less than 0.0001 Pa at 20 °C 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 expected to be significant. However, the substance may be available for respiratory absorption in the lung after inhalation of aerosols, if the substance is sprayed. In humans, particles with aerodynamic diameters below 100μm have the potential to be inhaled. Particles with aerodynamic diameters below 50μm may reach the thoracic region and those below 15μm the alveolar region of the respiratory tract. Particles deposited in the nasopharyngeal/thoracic region will mainly be cleared from the airways by the mucocilliary mechanism and swallowed (ECHA, 2017).

 

Lipophilic compounds with a log Pow > 4, that are poorly soluble in water (1 mg/L or less) can be taken up by micellar solubilisation. Esterases present in the lung lining fluid may also hydrolyse the substance, hence making the resulting alcohol and fatty acids available for respiratory absorption. Due to the high molecular weight of the parent substance, absorption is driven by enzymatic hydrolysis of the ester to the respective metabolites and subsequent absorption. However, as discussed above, hydrolysis of fatty acid esters with more than 3 ester bonds is considered to be slow (Mattson und Volpenhein, 1972a) and the possibility of the test substance to be hydrolysed enzymatically to the respective metabolites and their subsequent absorption is considered to be low as well.

 

In addition, available data on toxicity after inhalation of aerosols of the structural related substances Pentaerythritol tetraesters of n-decanoic, n-heptanoic, n-octanoic and n-valeric acids (CAS No. 68424-31-7; Zeneca, 1994a), Fatty acids, C5-9 tetraesters with pentaerythritol (CAS No. 67762-53-2; Mobil, 1990; Huntington, 1999b), and Fatty acids, C5-9, mixed esters with dipentaerythritol and pentaerythritol (CAS No. 85536-35-2; Zeneca, 1994b) are also considered for assessment of inhalation absorption. In acute inhalation toxicity studies conducted with those source substances, no signs of systemic toxicity were observed. The inhalation LC50 values determined range from > 4.06 mg/L air to > 5 mg/L air for male and female rats and correspond to the highest concentrations tested in each study. Moreover, in a subchronic (90-day) repeated dose toxicity study performed with the Fatty acids, C5-9, tetraesters with pentaerythritol (CAS No. 67762-53-2), no toxicologically relevant effects were observed up to and including the highest dose level of 0.5 mg/mL air in male and female rats (Mobil, 1992). The data indicate that respiratory absorption of the test substances is not higher than absorption through the intestinal epithelium as no systemic effects have been observed in any inhalation study. However, the lack of systemic toxicity might also be attributed to no or a low toxicity. It can therefore be taken only as indication but not as hard proof of a low absorption after inhalation exposure.

 

Overall, a systemic bioavailability of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids in humans is not considered likely after inhalation. The absorption rate is not expected to be higher than that following oral exposure as no systemic toxicity was observed with structural analogue substances after inhalation exposure. The lack of inhalation toxicity hints that respiratory absorption is limited. It is important to note that due to the low vapour pressure, exposure via the inhalation route is expected only if aerosols or droplets of an inhalable size (i.e. < 15 µm) are generated, e.g. in spray applications.

 

Accumulation and distribution

Distribution of a compound within the body through the circulatory system depends on the physico-chemical 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 its extracellular concentration, particularly in fatty tissues. Furthermore, the concentration of a substance in blood or plasma and subsequently its distribution depends on the rates of absorption. Although there is no direct correlation between the lipophilicity of a substance and its biological half-life, it is generally accepted that substances with high log Pow values have long biological half-lives. The high log Pow > 6.5 of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids is therefore indicative of the potential to accumulate in adipose tissue (ECHA, 2017). However as the absorption of the substance is considered to be very low, the potential of its bioaccumulation is very low as well.

 

As discussed in length under oral absorption, esters of PE and DiPE and fatty acids will undergo slow esterase-catalysed hydrolysis, leading to the hydrolysis products PE, DiPE and the fatty acids. Therefore, an assessment of distribution and accumulation of the hydrolysis products is considered more relevant. Pentaerythritol (PE) and dipentaerythritol (DiPE) are highly soluble in water (PE: 25 mg/L; log Pow < 0.3; OECD SIDS, 1998; DiPE: 3000 mg/L, log Pow = -2; OECD SIDS, 1995) with a low molecular weight. They will be distributed in aqueous fluids by diffusion through aqueous channels and pores. There is no protein binding assumed and they are distributed poorly in fatty tissues. Consequently, there is no potential to accumulate in adipose tissue. After being absorbed, fatty acids, are (re-)esterified along with other fatty acids into triglycerides and released in chylomicrons into the lymphatic system. Fatty acids of carbon chain length ≤ 12 may be transported directly to the liver via the portal vein as the free acid bound to albumin, instead of being re-esterified. Chylomicrons are transported in the lymph to the thoracic duct and subsequently to the venous system. On contact with the capillaries, enzymatic hydrolysis of chylomicron triacylglycerol fatty acids by lipoprotein lipase takes place. Most of the resulting fatty acids are taken up by adipose tissue and re-esterified into triglycerides for storage. Triacylglycerol fatty acids are likewise taken up by muscle and oxidized to derive energy or they are released into the systemic circulation and returned to the liver, where they are metabolised, stored or re-enter the circulation (IOM, 2005; Johnson, 1990; Johnson, 2001; Lehninger, 1998; NTP, 1994; Stryer, 1996; WHO, 2001). There is a continuous turnover of stored fatty acids, as they are constantly metabolised to generate energy and then excreted as CO₂. Accumulation of fatty acids takes place only if their intake exceeds the caloric requirements of the organism.

 

Therefore, the available information indicates that no significant bioaccumulation in adipose tissue of the parent substance and its hydrolysis products is anticipated.

 

Metabolism

Esters of fatty acids are hydrolysed to the corresponding alcohol and fatty acid by esterases (Fukami and Yokoi, 2012). Depending on the route of exposure, esterase-catalysed hydrolysis takes place at different places in the organism. After oral ingestion, esters are hydrolised already in the gastro-intestinal fluids. In contrast, esters which are absorbed through the pulmonary alveolar membrane or through the skin enter the systemic circulation directly before they are transported to the liver where hydrolysis will basically take place. Thus, Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids is expected to be hydrolysed to pentaerythritol, dipentaerythritol and the respective fatty acids by esterases, even though the hydrolysis rate is expected to be low. The hydrolysis of fatty acid esters containing more than 3 ester groups is assumed to be slow as already discussed above. In vivo studies in rats demonstrated a decrease in absorption with increasing esterification grade. For example, for Pentaerythritol tetraoleate an absorption rate of 64% and 90% (25% and 10% of dietary fat) was observed, respectively, while an absorption rate of 100% was observed for Glycerol trioleate when ingested at 100% of dietary fat (Mattson and Nolen, 1972). In addition it has been shown in vitro that the hydrolysis rate of pentaerythritol tetraoleate was lower when compared with the hydrolysis rate of the triglyceride Glycerol trioleate (Mattson and Volpenhein, 1972a).

 

Pentaerythritol (PE) is absorbed rapidly but excreted unchanged. DiCarlo et al. (1965) reported that 10 mg/kg 14C-labled PE orally administered to mice was absorbed and excreted rapidly from the GIT. Almost half of the administered dose left the GIT within 15 minutes and 68% of the dose appeared as unchanged PE in the urine and faeces after 4 hours already. A similar metabolic behaviour can be expected for DiPE. Fatty acids are degraded by mitochondrialβ-oxidation which takes place in most animal tissues and uses an enzyme complex for a series of oxidation and hydration reactions, resulting in the cleavage of acetate groups in the form of acetyl-CoA. The alkyl chain length is reduced by 2 carbon atoms during eachβ-oxidation cycle. Alternative pathways for oxidation can be found in the liver (ω-oxidation) and the brain (α-oxidation). Iso-fatty acids such as isooctadecanoic acid have been found to be activated by acyl coenzyme A synthetase of rat liver homogenates and to be metabolised to a large extent byω-oxidation. Each C2-unit resulting fromβ-oxidation enters the citric acid cycle as acetyl-CoA, through which they are completely oxidized to CO₂ (CIR, 1983, 1987; IOM, 2005; Lehninger, 1998; Stryer, 1996; WHO, 2001; Matulka, 2009).

 

The potential metabolites following enzymatic degradation of the test substance were predicted using the QSAR OECD Toolbox (OECD, 2017). This QSAR tool predicts the primary and secondary metabolites of the parent compound that may result from enzymatic activity in the liver, in the skin and by the micro-flora in the GIT. 11 to 14 hepatic metabolites and 2 dermal metabolites were predicted for the low molecular weight constituent and the high molecular weight constituent of Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids. As can be expected, the typical metabolic transformation of the parent compound is the cleavage of the various ester bonds that takes place mainly in the liver. Metabolites formed in the skin are identified by addition of a hydroxyl group to the alkyl chain of the esters. Following the first reaction step, hydrolysis products may be metabolised further. The resulting liver and skin metabolites are the products ofα-,β- orω-oxidation (i.e. addition of a hydroxyl group). In the case ofω-oxidation, it is followed by further oxidation to the aldehyde, which is then oxidised to the corresponding carboxylic acid. The ester bond may also remain intact, in which case a hydroxyl group is added to an alkyl chain. In general, the hydroxyl groups make the substances more water-soluble and susceptible to metabolism by phase II enzymes. The metabolites formed in the skin are expected to enter the blood circulation and have the same fate as the hepatic metabolites. Between 23 and 65 metabolites were predicted to result from all kinds of microbiological metabolism for the low molecular weight and high molecular weight constituents, respectively. This rather high number includes many minor variations in the C-chain length and number of carbonyl and hydroxyl groups, reflecting the diversity of many microbial enzymes identified. Not all of these reactions are expected to take place in the human GIT. The results of the OECD Toolbox simulation support the information on metabolism routes retrieved in the literature.

 

Excretion

On the basis of its low absorption, one route of excretion for Monopentaerythritol tetraesters and dipentaerythritol hexaesters of valeric, heptanoic and nonanoic acids is expected to be by biliary excretion via faeces. Hydrolysis of the parent compound yields fatty acids, PE and DiPE. The fatty acids will be metabolised for energy generation or stored as lipids in adipose tissue or used for further physiological processes, e.g. incorporation into cell membranes (Lehninger, 1970; Stryer, 1996). Therefore, the fatty acid components are not expected to be excreted to a significant degree via the urine or faeces but excreted via exhaled air as CO2 or stored as described above. PE is not metabolized but excreted unchanged via urine. 10 mg/kg 14C-labled PE orally administered to mice was absorbed from the gastrointestinal tract rapidly and excreted via urine. Almost half of the administered dose left the gastrointestinal tract within 15 minutes and 68% of the dose appeared as unchanged PE in the urine and faeces after 4 hours (DiCarlo et al., 1965). The amount found in faeces was assumed to be contamination from urine due to the setup of the metabolic cages. Additionally, Kutscher (1948) found 85 - 87% of unaltered PE in the urine of humans ingesting PE. Since dipentaerythritol is even more water soluble, a similar excretion scheme is expected to be applicable.

 

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