Fatty acids, C8-10, mixed esters with neopentyl glycol and trimethylolpropane

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29 Aug - 30 Nov 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions (no data on GLP and analytical purity, no hydrolysis test in saliva and gastric juice simulants performed, limited details in reporting).

Objective of study:

other: hydrolysis in intestinal fluid simulant

Qualifier:

equivalent or similar to guideline

Guideline:

other: EFSA Note for Guidance for Food Contact Materials Annex 1 to Chapter III MEASUREMENT OF HYDROLYSIS OF PLASTICS MONOMERS AND ADDITIVES IN DIGESTIVE FLUID SIMULANTS

Deviations:

yes

Remarks:

no hydrolysis test in saliva and gastric juice simulants; limited details in reporting

GLP compliance:

not specified

Radiolabelling:

no

Species:

other: not specified; presumably pig in accordance with the test method used

Strain:

not specified

Sex:

not specified

Details on test animals or test system and environmental conditions:

TEST DIGESTIVE SIMULANTS
INTESTINAL FLUID SIMULANT
- Description: intestinal fluid simulant according to “Note of Guidance for Food Contact Materials”, no further details given.

Route of administration:

other: mixing

Vehicle:

other: acetonitrile

Details on exposure:

HYDROLYSIS WITH INTESTINAL-FLUID SIMULANT:
For the hydrolysis investigation, the esters were dissolved in acetonitrile. These solutions were added to the intestinal-fluid simulant tempered to 37 °C. The concentration of acetonitrile in the reaction mixture was about 0.1%. Samples were taken after 0, 1, 2 and 4 h.

Duration and frequency of treatment / exposure:

0, 1, 2 and 4 h

Dose / conc.:

22.77 ppm

No. of animals per sex per dose / concentration:

not applicable, the test was performed in triplicates

Control animals:

other: not applicable

Details on dosing and sampling:

DETERMINATION OF HYDROLYSIS PRODUCTS
- Principle: following incubation, a naphthalene solution (solved in acetone) was added as an internal standard to the samples. Afterwards, the enzymes were precipitated by the addition of ice-cold acetone. After filtration the acetone was evaporated. The aqueous solutions were acidified with 0.1 M hydrochloric acid (pH 1.2) and were extracted 3 times with dichloromethane. After addition of an alkane standard (tridecane) and derivatisation with N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA) at 60°C for 1 h the concentrated dichloromethane solutions were analysed by gas chromatography coupled with a mass spectrometer (GC/MS). Quantification of the esters and the hydrolysis products was performed specifically by external calibration curves.
- Recovery assays: a duplicate of 3 different concentrations of the acid (hydrolysis product) were performed. For the recovery investigations the acid was dissolved in acetonitrile. These solutions were added to the intestinal-fluid simulant tempered to 37°C. After 4 h a naphthalene solution in acetone was added as an internal standard to the samples and the enzyme was precipitated by the addition of ice-cold acetone. Work-up and quantification was performed as described above.
Recovery of the ester was determined using a hydrolysis sample analogue to the "0 hour" assay.

Statistics:

Mean values of triplicates were calculated.

Type:

other: ester hydrolysis in intestinal fluid simulant

Results:

85.7, 86.1 and 89.4% after 1, 2 and 4 h, respectively

Details on absorption:

not applicable

Details on distribution in tissues:

not applicable

Details on excretion:

not applicable

Metabolites identified:

yes

Details on metabolites:

Quantification of the parent substance 2,2-dimethyl-1,3-propandiolheptanoate after hydrolysis in intestinal-fluid simulant after 0, 1, 2 and 4 h hydrolysis, respectively:
22.77, 3.26, 3.17 and 2.41 ppm corresponding to 100, 14.3, 13.9 and 10.6% of the initial test concentration. Thus, 2,2-dimethyl-1,3-propandiolheptanoate was hydrolysed to nearly 90% after pancreatic digestion for 4 h.

Quantification of the free fatty acid after hydrolysis in intestinal-fluid simulant after 0, 1, 2 and 4 h hydrolysis, respectively:
0, 15.72, 15.86 and 17.06 ppm

Table 1. Hydrolysis of 2,2 -dimethyl-1,3-propandiolheptanoate with intestinal-fluid simulant

Contact time (h)	Results
	Ester (ppm)	Ester (%)	Acid (ppm)
0	22.77	100.0	0.000
1	3.26	14.3	15.72
2	3.17	13.9	15.86
4	2.41	10.6	17.06

Table 2. Mass balance of the ester hydrolysis

Contact time (h)	Results
	Ester (µmol)	Acid (µmol) (calc.)	Acid (µmol) (exp.)
0	0.347	0.000	0.000
1	0.050	0.594	0.604
2	0.048	0.597	0.609
4	0.037	0.620	0.655

Table 3. Recoveries of ester and acid

Analyte	Results
	ppm (calc.)	ppm (exp.)	Recovery (%)
Ester (from pancreatic medium)	22.77	5.43	23.9
Ester (from water)	13.90	13.12	94.4
	30.88	31.29	101.3
	46.32	51.52	111.2
Acid (from pancreatic medium)	12.82	9.28	72.4
	28.48	23.18	81.4
	49.84	36.19	72.6

Conclusions:

bioaccumulation potential cannot be judged based on study results

Reference 2

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

Please refer to the Additional Information field in the endpoint study summary

Reason / purpose for cross-reference:

read-across source

Type:

other: ester hydrolysis in intestinal fluid simulant

Results:

85.7, 86.1 and 89.4% after 1, 2 and 4 h, respectively (Croda, 2012)

Metabolites identified:

yes

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Additional information

Basic toxicokinetics

There are no experimental studies available in which the toxicokinetic behaviour of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) has been investigated.

 

Therefore, in accordance with Annex VIII, Column 1, Item 8.8.1, of Regulation (EC) No 1907/2006 and with Guidance on information requirements and chemical safety assessment Chapter R.7c: Endpoint specific guidance (ECHA, 2014), an assessment of the toxicokinetic behaviour of the substance Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) was conducted to the extent that can be derived from relevant available information on physico-chemical and toxicological properties and taking into account available information on polyol esters.

 

The substance Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) mainly consists of diester of C8 and C10 saturated fatty acids with 2,2-dimethyl-1,3-propanediol (neopentylglycol, NPG) and triester of C8 and C10 saturated fatty acids with 1,1,1-trimethylolpropan (TMP) which meets the definition of an UVCB substance based on the analytical characterization. The substance is an organic liquid which is poorly water soluble <0.15 mg/L at 20 °C and pH=6.3-6.5 (BASF, 2012), with a molecular weight range of 356.54 - 412.65g/mol for the C8/C10 diester with NPG and 512.78 - 596.94 g/mol for the C8/C10 triester with TMP, log Pow of 7.66 - 13.59 (KOWWIN v1.68) and a vapour pressure <0.00001 Pa at 20 °C (QSAR, SPARC v4.6) for C8/C10 diester with NPG and C8/C10 triester with TMP.

 

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, 2014).

 

Oral

The smaller the molecule, the more easily it will be taken up. In general, molecular weights below 500 are favourable for oral absorption (ECHA, 2014). With a molecular weight range of 356.54 - 596.94, absorption of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) is in general expected to be low in the gastrointestinal (GI) tract. The predicted log Pow >4 suggests that Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) may be absorbed by micellar solubilisation, as this mechanism is of importance for highly lipophilic substances that are poorly soluble in water (1 mg/L or less).

 

Applying the “Lipinski Rule of Five” (Lipinski et al. (2001), refined by Ghose et al. (1999)) to Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8), shows that two rules are not fulfilled: the substance has a molecular weight range above 500 and has more than 10 hydrogen bond. This indicates a moderate to low potential for oral absorption.

 

Metabolism of the test substance via enzymes and the microflora in the gastrointestinal (GI) tract may occur. In fact, after oral ingestion, fatty acid esters with glycerol (glycerides) are rapidly hydrolysed by ubiquitously expressed esterases (Mattsson and Volpenhein, 1972a). In general, it is assumed that the hydrolysis rate varies depending on the fatty acids/alcohol combinations, and grade of esterification (Mattson and Volpenhein, 1969; Mattson and Volpenhein, 1972a,b). For polyol esters, a lower rate of enzymatic hydrolysis in the GI tract was shown for compounds with more than 3 ester groups (Mattson and Volpenhein, 1972a, b). The in vitro hydrolysis rate of pentaerythritol ester was about 2000 times slower in comparison to glycerol esters (Mattson and Volpenhein, 1972a, b). Based on the available literature Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8), a diester/triester of C8 and C10 fatty acids and NPG and TMP, is considered to undergo stepwise chemical changes in the GI-fluids as a result of enzymatic hydrolysis after oral ingestion. Furthermore, the result of the pancreatic digestion of the structurally related substance heptanoic acid, ester with 2,2-dimethyl-1,3 –propandiol, shows a degradation of the ester of almost 90% within 4 hours (Croda, 2012a).

 

The physico-chemical characteristics of the cleavage products (e.g. physical form, water solubility, molecular weight, log Pow, vapour pressure) are likely to be different from those of the parent substance, and hence the predictions regarding absorption based on the physico-chemical characteristics of the parent substance no longer apply (ECHA, 2014). The cleavage products are anticipated to be absorbed in the GI tract. The highly lipophilic fatty acids (covering C8 to C10 saturated fatty acids) are expected be absorbed by micellar solubilisation (Ramirez et al., 2001), and the alcohol moiety (NPG/TMP) is readily dissolved into the GI-fluids and will be absorbed because of its physico-chemical parameters (MW = 104.15 g/mol, log Pow = 0.12 at 25 °C and moderate water solubility (OECD SIDS, 1993) for NPG and MW = 134.20 g/mol, log Pow = -0.47 at 25 °C and high water solubility >100 g/L at 20 - 25 °C for TMP (OECD SIDS, 1994)).

 

The indications that the target substance has relatively low oral absorption and/or low acute toxicity due to its physico-chemical characteristics are supported by the available data on acute oral toxicity for Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) with a LD50 value >2000 mg/kg bw (BASF, 1988a).

 

Overall, systemic bioavailability of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) and/or the respective cleavage products in humans is considered likely after oral uptake of the substance.

 

Dermal

The smaller the molecule, the more easily it may be taken up. In general, a molecular weight below 100 favours dermal absorption, above 500 the molecule may be too large (ECHA, 2014). If the substance is a skin irritant or corrosive, damage to the skin surface may enhance penetration (ECHA, 2014). As the results with the target and source substances do not indicate skin irritating potential, enhanced penetration of the substance due to local skin damage can be excluded.

 

This assumption is supported by the physico-chemical properties of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8): 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 hence limit absorption across the skin. Therefore, the uptake of such substances into the stratum corneum itself is limited (ECHA, 2014). As the molecular weight of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) is 356.54 - 596.94 g/mol, water solubility is less than 1 mg/L and the log Pow >4, dermal uptake of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) is expected to be low.

 

Additionally, QSAR based dermal permeability prediction (EPA, 2012) using molecular weight, log Pow and water solubility was performed resulting in a dermal penetration rate ranging from 2.61 (diester of C8 saturated fatty acids with NPG) to 4820 µg/cm²/h (triester of C10 saturated fatty acids with TMP) for the target substance Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8). This values are in contrast to the general assumption described above and therefore considered as an indicator for a medium (40%) to high (80%) dermal absorption rate for Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8).

 

Overall, the calculated medium to high dermal absorption potential, the low water solubility, the molecular weight (>100), the high log Pow value and the fact that the substance is not irritating to skin implies that dermal uptake of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) in humans is likely.

 

Inhalation

Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) has a low vapour pressure of <0.00001 Pa at 20 °C (QSAR, SPARC v4.6) and therefore 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 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, 2014). Lipophilic compounds with a log Kow >4 that are poorly soluble in water (1 mg/L or less) like Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) can be taken up by micellar solubilisation.

 

An acute inhalation toxicity study was performed with the structurally related substance heptanoic acid, ester with 2,2-dimethyl-1,3-propanediol (CAS 68855-18-5), in which rats were exposed nose-only to up to 5.22 mg/L of an aerosol for 4 hours (Croda, 2012b). No mortality occurred and no toxicologically relevant effects were observed at the end of the study period.

 

Overall, systemic bioavailability of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) in humans is considered likely after inhalation of aerosols but is not expected to be higher than following oral exposure.

 

Accumulation

Highly lipophilic substances tend in general to concentrate in adipose tissue, and depending on the conditions of exposure may accumulate within the body. Although there is no direct correlation between the lipophilicity of a substance and its biological half-life, it is generally the case that substances with high log Pow values have long biological half-lives. The high log Pow of >7 implies that Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) may have the potential to accumulate in adipose tissue (ECHA, 2014).

 

However, as further described in the section ‘Metabolism’ below, esters of alcohols and fatty acids undergo esterase-catalysed hydrolysis, depending of the fatty acid chain length and degree of esterification, leading to the respective cleavage products, namely fatty acids and alcohol moieties.

 

The alcohol moiety NPG and TMP are the first cleavage products of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8). Due to its physico-chemical properties (high water solubility and low molecular weight), accumulation of NPG and TMP in adipose tissue is considered to be unlikely. The second cleavage products, the fatty acid moieties (C8 and C10), can be stored as triglycerides in adipose tissue or be incorporated into cell membranes. At the same time, fatty acids are also required as a source of energy. Therefore, stored fatty acids underlie a continuous turnover as they are permanently metabolized and excreted. Bioaccumulation of fatty acids only takes place if their intake exceeds the caloric requirements of the organism.

 

Overall, the available information indicates that no significant bioaccumulation in adipose tissue of the parent substance and cleavage products will occur.

 

Distribution

Distribution within the body through the circulatory system depends on the molecular weight, the lipophilic character and water solubility of a substance. 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, 2014).

 

The enzymatic hydrolysis of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) will result in the cleavage products NPG, TMP and the respective fatty acids. NPG and TMP will be distributed in aqueous compartments of the organism and may also be taken up by various tissues. Similarly, fatty acids are also distributed in the organism and can be taken up by various tissues. They can be stored as triglycerides in adipose tissue depots or be incorporated into cell membranes (Masoro, 1977).

 

Overall, the available information indicates that the cleavage products, NPG, TMP and fatty acids will be distributed in the organism.

 

Metabolism

Esters of fatty acids are hydrolysed to the corresponding alcohol and fatty acids by esterases (Fukami and Yokoi, 2012). Therefore, Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) is expected to undergo hydrolysis within the human body. 

Depending on the route of exposure, esterase-catalysed hydrolysis takes place at different places in the organism: after oral ingestion, esters of alcohols and fatty acids undergo enzymatic hydrolysis already in the GI fluids. In contrast, substances which are absorbed through the pulmonary alveolar membrane or through the skin enter the systemic circulation directly before entering the liver where hydrolysis will take place. The first cleavage products, NPG and TMP, are likely to be conjugated by UGTs. The glucuronidated products are then excreted in the urine (Gessner, 1960). The other cleavage products, C8 and C10 saturated fatty acids, are metabolised by stepwise beta-oxidation before entering the citric acid cycle as acetyl CoA (Stryer 1996).

 

Excretion

No data on excretion of Fatty acids, C8-10 (even numbered), diesters with neopentyl glycol and di- and triesters with trimethylolpropane (CAS 97281-24-8) is available. Based on the anticipated enzymatic hydrolysis, fatty acids, NPG and TMP as breakdown products will be present in the body. The fatty acid components will be metabolized for energy generation and afterwards mainly excreted by expired air as CO2, or stored as lipids in adipose tissue or used for further physiological properties e.g., incorporation into cell membranes (Stryer, 1996). Therefore, the C8 and C10 saturated 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. The main route for elimination of the alcohol is renal excretion via urine (Gessner, 1960).

References

ECHA (2014). Guidance on information requirements and chemical safety assessment, Chapter R.7c: Endpoint specific guidance.

EPA (2012). Dermwin v2.02, Estimation Programs Interface Suite™ for Microsoft® Windows, v 4.11. United States Environmental Protection Agency, Washington, DC, USA. Downloaded fromhttp://www.epa.gov/opptintr/exposure/pubs/episuitedl.htm.

Fukami, T. and Yokoi, T. (2012). The Emerging Role of Human Esterases. Drug Metabolism and Pharmacokinetics, Advance publication July 17th, 2012.

Gessner PK, Parke DV, Williams RT (1960) Studies in detoxication. 80. The metabolism of glycols Biochem J 74: 1-5.

Ghose et al. (1999). A Knowledge-Based Approach in Designing Combinatorial or Medicinal Chemistry Libraries for Drug Discovery. J. Comb. Chem. 1 (1): 55-68.Lehninger, A.L. (1970). Biochemistry. Worth Publishers, Inc.

OECD SIDS (1993)http://www.inchem.org/documents/sids/sids/126307.pdf

OECD SIDS (1994)www.inchem.org/documents/sids/sids/77996.pdf

Lipinski et al. (2001). Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Del. Rev. 46: 3-26.

Masoro (1977). Lipids and lipid metabolism. Ann. Rev. Physiol.39: 301-321.

Mattson, F.H. and Volpenhein, R.A. (1969): Relative rates of hydrolysis by rat pancreatic lipase of esters of C2 - C18 fatty acids with C1 – C18 primary n-alcohols. J Lipid Res Vol(10): 271 – 276.

Mattson F.H. and Volpenhein R.A., (1972a): Hydrolysis of fully esterified alcohols containing from one to eight hydroxyl groups by the lipolytic enzymes of rat pancreatic juice. J Lip Res 13, 325-328

Mattson F.H. and Volpenhein R.A., (1972b): Digestion in vitro of erythritol esters by rat pancreatic juice enzymes. J Lip Res 13, 777-782

Ramirez et al. (2001). Absorption and distribution of dietary fatty acids from different sources. Early Human Development 65 Suppl.: S95–S101.

Stryer, L. (1996): Biochemie. 2nd revised reprint, Heidelberg; Berlin; Oxford: Spektrum Akad. Verlag.