Glycerides, C16-18 (even numbered) and their amidation products with diethylenetriamine

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1996

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Justification for type of information:

Pharmacokinetics and Material Balance Studies of Diethylenetriamine Trihydrochloride in the Fischer 344 Rat is presented due to potential release of Diethylenetriamine from the registered substance due to metabolic activation and presence of small quantities within the Amidoamine and Amidoamine 2 substances.

Objective of study:

toxicokinetics

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Deviations:

yes

Principles of method if other than guideline:

Pharmacokinetics and Material Balance study of Diethylenetriamine Trihydrochloride in the Fischer 344 comparable to OECD 417 requirements.

GLP compliance:

not specified

Specific details on test material used for the study:

[1,2-14C]-Diethylenetriamine trihydrochloride (14C-DETA·3HCl) with a specific activity of 6 mCi mmol−1 and a radiochemical purity of 98% (CAS 3488-89-9).

Radiolabelling:

yes

Species:

rat

Strain:

Fischer 344

Details on species / strain selection:

Male Fischer 344 rats, 36 days old, were obtained from Charles River Breeding Laboratories, Portage MI. After a 9–14-day acclimation period, healthy animals were assigned to study groups based on a body-weight-stratified randomization procedure. Throughout the study rats were maintained on a 12-h light/dark cycle under controlled temperature and humidity conditions, and food and water were available ad libitum.

Sex:

male

Details on test animals or test system and environmental conditions:

Male Fischer 344 rats, 36 days old, were obtained from Charles River Breeding Laboratories, Portage, MI. After a 9–14-day acclimation period, healthy animals were assigned to study groups based on a body-weight-stratified randomization procedure. Throughout the study rats were maintained on a 12-h light/dark cycle under controlled temperature and humidity conditions, and food and water were available ad libitum.

Route of administration:

other: Endotracheal, Intravenous and Oral

Vehicle:

other: unlabeled DETA and physiological saline

Duration and frequency of treatment / exposure:

Single dosing of animals

Dose / conc.:

50 mg/kg bw/day (nominal)

Remarks:

Intravenous, oral and endotracheal route

Dose / conc.:

500 mg/kg bw/day (nominal)

Remarks:

Oral and endotracheal route

No. of animals per sex per dose / concentration:

Dose route: Intravenous/Oral/Endotracheal
Number of rats: 4/5/4

Control animals:

no

Details on dosing and sampling:

Dosing solutions were prepared by mixing appropriate amounts of unlabeled DETA with [14C]-DETA·3HCl in physiological saline to provide target dosages of 50 and 500 mg kg−1 at a constant dosing volume of 0.25 ml per animal.
The dosing solutions were adjusted to pH 7.4 using hydrochloric acid or ammonium hydroxide, as required. Animals in the pharmacokinetics study groups had an indwelling cannula implanted in the right jugular vein1 day prior to dosing. Intravenous dosing was accomplished by injecting
the tail vein. Oral dosing was performed with a stainless-steel feeding needle, and endotracheal instillation was achieved by injecting through a small polyethylene tube into the trachea of rats under light anesthesia with Metofane.

For the material balance study, the animals were placed in all-glass Roth metabolism cages immediately after dosing. Expired 14CO2 was trapped using a solution of 2-methoxyethanol–ethanolamine (7:3, v/v) and sampled for radioactivity 6, 24 and 48 h after dosing. Urine and feces were collected 24 and 48 h after dose administration. Cages were washed with a 50% aqueous acetone solution and samples of the wash solution
were taken for radiometric analysis. Blood was obtained from the abdominal aorta 48 h after dosing. Animals were killed by exsanguination after collection of the last blood and excreta samples. Selected tissues were removed, sampled and analyzed for radioactivity. The remaining carcasses were solubilized in 10 N NaOH and samples of this solution were also analyzed for radioactivity. For the pharmacokinetics study, rats
were placed in stainless-steel wire metabolism cages. Blood samples (about 0.2 ml) were taken via the jugular cannulae at 5, 15, 30 and 45 min and at 1, 2, 3, 4, 5, 6, 8, 10, 12, 16 and 24 h after dosing.

The amount of radioactivity in various samples was measured by liquid scintillation spectrometry. Blood was centrifuged in heparinized capillary tubes to separate into plasma and packed red cells. Portions of other liquid samples, such as plasma, urine and cage washes, were weighed directly into scintillation vials containing 10 ml of Aquasol (New England Nuclear, Boston, MA). Samples of the CO2 trapping solution and solubilized carcass were weighed directly into scintillation vials and counted in a solution of equal volumes of 2-methoxyethanol, dioxane and xylene containing 160:120:1 (w/w/w) of naphthalene, 2,5-diphenyloxazole and 2,2-p-phenylenebis-(5-phenyloxazole). Feces and tissues were solubilized in Soluene 350 (Packard Instrument Co., Downers Grove, IL), oxidized with 30% H2O2 and counted with 10 ml of Dimilume 30 (Packard Instrument Co., Downers Grove, IL).

Statistics:

Curve fitting of the pharmacokinetic data was performed by hand feathering or with the aid of ESTRIP. Parameters estimated included area under the concentration–time curve to infinite time (AUC`), total clearance and terminal half-lives. The maximal plasma concentration (Cmax) and time to maximal plasma concentration (tCmax) following oral or endotracheal dosing were estimated from the concentration–time course equation by the Newton–Raphson method of numerical analysis.

Type:

absorption

Results:

The bioavailability was over 90%. [14C]-DETA·3HCl was well absorbed following oral or endotracheal dosing. Clearance of absorbed [14C]- DETA·CHCl from the plasma was also rapid, with a terminal half-life of 9-16 h.

Type:

distribution

Results:

The highest radioactivity concentrations of DETA was attained in the liver and kidney, suggesting that these organs are common targets for this alkyleneamine

Type:

metabolism

Results:

Metabolism of [14C]-DETA·3HCl appeared to play a small role in the elimination of DETA from the body, as unchanged DETA was the principal component excreted in the urine.

Type:

excretion

Results:

The major excretory route of DETA was via the urine and feces. These two pathways together accounted for over 70% of the total dose excreted.

Details on distribution in tissues:

Material balance of radioactivity in Fischer 344 rats at 48 h following an oral or endotracheal dose of [14C]-diethylenetriamine trihydrochloride

Dose route Oral Endotracheal
Targeted dosage (mg kg-1) 50 500 50 500
Body weight (kg) 0.169 ± 0.006 0.202 ± 0.013 0.171 ± 0.007 0.202 ± 0.004
Radioactivity (mCi) 7.5 ± 0.3 10.2 ± 0.2 7.8 ± 0.15 9.6 ± 0.10
Dose (mg) 8.2 ± 0.3 102.8 ± 2.5 8.5 ± 0.1 97.2 ± 0.9
Dosage (mg kg-1) 48.2 ± 2.7 511 ± 29.5 49.5 ± 2.0 481 ± 5.5
(Per cent of dosed radioactivity)

Urine 31.1 ± 2.5 42.9 ± 9.3 32.2 ± 9.2 40.3 ± 9.5
Feces 45.6 ± 2.1 44.2 ± 1.3 39.7 ± 8.7 45.5 ± 8.4
CO2 1.09 ± 0.10 0.52 ± 0.04 1.28 ± 0.18 0.57 ± 0.12
Cage washings 5.4 ± 4.9 10.1 ± 5.6 8.0 ± 2.6 14.1 ± 6.2
Carcass 2.0 ± 0.4 1.8 ± 0.1 3.1 ± 0.6 2.1 ± 0.4
Total recovery 85.2 ± 4.3 99.6 ± 3.3 84.3 ± 4.4 102.8 ± 1.2

Key result

Test no.:

#1

Toxicokinetic parameters:

AUC: 62.2 ± 16.9 mg·h ml-1

Remarks:

intravenous route

Key result

Test no.:

#2

Toxicokinetic parameters:

AUC: 59.1 ± 4.1 mg·h ml-1

Remarks:

oral route

Key result

Test no.:

#3

Toxicokinetic parameters:

AUC: 56.0 ± 9.1 mg·h ml-1

Remarks:

endotracheal route

Key result

Test no.:

#1

Toxicokinetic parameters:

Cmax: 120 ± 21 mg ml-1

Remarks:

intravenous route

Key result

Test no.:

#2

Toxicokinetic parameters:

Cmax: 13.83 ± 3.97 mg ml-1

Remarks:

oral route

Key result

Test no.:

#3

Toxicokinetic parameters:

Cmax: 10.83± 3.62 mg ml-1

Remarks:

endotracheal route

Metabolites identified:

yes

Remarks:

Urinary radioactivity was measured by cation exchange chromatography. The cation exchange column was able to resolve unchanged DETA. The neutral fraction probably probably consisted of a deaminated metabolite of DETA.

Details on metabolites:

not reported

Enzymatic activity measured:

not reported

Bioaccessibility (or Bioavailability) testing results:

Bioavailability (oral): 95 %
Bioavailability (endotracheal): 90 %

DETA·3HCl-derived radioactivity concentrations in the plasma could best be described by a tri-exponential equation of the form C = 119.6e−8.49t + 69.4e−2.07t + 2.4e−0.12t . The maximal plasma concentration (Cmax) was 120 ± 21 mg ml−1 and the apparent volume of distribution (Vd) was 486 ± 220 ml kg−1. In rats dosed orally or endotracheally, the peak concentration was generally attained within 1 h following dosing , suggesting that absorption of [14C]- DETA·3HCl from the gut or the respiratory tract was quite rapid. The bioavailability was over 90%, suggesting that [14C]-DETA·3HCl was well absorbed following oral or endotracheal dosing. Clearance of absorbed [14C]-DETA·CHCl from the plasma was also rapid, with a terminal half-life of 9–16 h. Fecal excretion was the major route of excretion, followed closely by elimination in the urine. Only a small percentage of the dose was recovered as expired CO2 or was retained in the tissues and carcasses. No significant differences in the percentages of radioactivity excreted in the urine, feces or CO2 were observed between animals dosed orally or endotracheally. The DETA was distributed throughout the body, with the highest concentrations found in the kidney and liver. The mean concentration for all tissues in the 500 mg kg−1 group was about 7.5 times that in the 50 mg kg−1 group. This was slightly lower than the 10-fold difference in dosages. Similar to the excretion data, there was no indication that route of administration had any significant effect on tissue concentrations.

 

Table 1. Dosages and pharmacokinetic parameters in Fischer 344 rats following intravenous, oral  and endotracheal dosing  with [14C]-diethylenetriamine trihydrochloride

Dose route	Intravenous	Oral	Endotracheal
Number of rats	4	5	4
Body weight (kg)	0.185 ± 0.005	0.178 ± 0.016	0.183 ± 0.002
Radioactivity (mCi)	48.7 ± 0.3	30.3 ± 0.4	30.6 ± 0.5
Dose (mg)	10.30 ± 0.08	8.33 ± 0.12	8.41 ± 0.14
Dosage (mg kg−1)	55.6 ± 1.9	46.8 ± 4.3	45.8 ± 1.2
Cmax (mg ml−1)	120 ± 21	13.83 ± 3.97	10.83± 3.62
tCmax (h)	-	0.77 ± 0.38	0.77 ± 0.36
Vd (ml kg−1)	486 ± 220	-	-
AUC` (mg·h ml−1)	62.2 ± 16.9	59.1 ± 4.1	56.0 ± 9.1
Bioavailability (%)	-	95.0	90.0
Total clearance (ml h−1)	174 ± 44	134 ± 10	138 ± 23
Terminal t1/2 (h)	9.7 ± 5.3	16.3 ± 6.1	9.0 ± 5.0

Table 2: Material balance of radioactivity in Fischer 344 rats at 48 h following an oral or endotracheal dose of [14C]-diethylenetriamine trihydrochloride

Dose route	Oral		Endotracheal
Targeted dosage (mg/kg)	50	500	50	500
Body weight (kg)	0.169 ± 0.006	0.202 ± 0.013	0.171 ± 0.007	0.202 ± 0.004
Radioactivity (mCi)	7.5 ± 0.3	10.2 ± 0.2	7.8 ± 0.15	9.6 ± 0.10
Dose (mg)	8.2 ± 0.3	102.8 ± 2.5	8.5 ± 0.1	97.2 ± 0.9
Dosage (mg/kg)	48.2 ± 2.7	511 ± 29.5	49.5 ± 2.0	481 ± 5.5
(Per cent of dosed radioactivity)
Urine	31.1 ± 2.5	42.9 ± 9.3	32.2 ± 9.2	40.3 ± 9.5
Feces	45.6 ± 2.1	44.2 ± 1.3	39.7 ± 8.7	45.5 ± 8.4
CO2	1.09 ± 0.10	0.52 ± 0.04	1.28 ± 0.18	0.57 ± 0.12
Cage washings	5.4 ± 4.9	10.1 ± 5.6	8.0 ± 2.6	14.1 ± 6.2
Carcass	2.0 ± 0.4	1.8 ± 0.1	3.1 ± 0.6	2.1 ± 0.4
Total recovery	85.2 ± 4.3	99.6 ± 3.3	84.3 ± 4.4	102.8 ± 1.2

 

Table 3. Distribution of radioactivity in tissues of Fischer 344 rats at 48 h following an oral or endotracheal dose of [14C]-diethylenetriamine trihydrochloride*

Dose route	Oral		Endotracheal
Dosage (mg/kg)	48	511	50	481
	(µg equivalent/g tissue)
Whole blood	0.9	5.3	0.9	5.9
Plasma	0.4	2.4	0.5	2.6
Blood cells	1.1	8.1	1.1	8.4
Adrenal	1.2	9.0	1.6	8.9
Bone marrow	1.0	8.6	1.7	8.7
Brain	0.2	1.1	0.2	1.1
Cecum	1.0	11.7	1.5	11.1
Esophagus	1.5	12.1	1.8	9.6
Fat	0.3	1.3	0.2	1.3
Gastrointestinal contents	0.4	7.0	1.1	27.4
Heart	0.9	5.4	1.1	5.4
Kidney	2.3	25.2	3.0	26.1
Large intestine	1.2	12.7	2.1	15.7
Liver	2.6	13.6	3.0	17.4
Lung	1.1	10.5	1.4	8.5
Lymph nodes	1.3	10.9	1.2	9.8
Muscle	0.6	4.8	0.7	4.6
Pancreas	0.6	3.4	0.7	5.2
Salivary glands	0.7	5.9	1.0	5.7
Skin	0.7	10.0	1.4	9.1
Spleen	1.1	10.8	1.8	11.0
Small intestine	2.1	5.3	0.8	5.9
Stomach	0.8	9.2	1.3	6.8
Testes	0.4	3.7	0.5	3.5
Thymus	1.2	11.8	1.8	11.1
Thyroid	1.1	6.6	1.4	7.3
Trachea	1.4	13.0	2.6	8.5
Urinary bladder	1.9	13.6	2.0	14.3

*Values are means of four animals.

Conclusions:

Metabolism of [14C]-DETA·3HCl appeared to play a small role in the elimination of DETA from the body, as unchanged DETA was the principal component excreted in the urine. The major excretory route of DETA was via the urine and feces. These two pathways together accounted for over 70% of the total dose excreted. The highest radioactivity concentrations of DETA was attained in the liver and kidney, suggesting that these organs are common targets for this alkyleneamine.

Relevance for the registered substance: The registered substance could potentially be hydrolized/metabolized to the extent that all amide bonds are cleaved enzymatically and DETA could be released. As major parts of DETA do not undergo any further metabolism in rats according to the results of this study liver and kidney could potential be target organs for adverse effects and eliminating the substance or potential metabolites.

Executive summary:

In the Klimisch 2 publication from Leung (1997) the pharmacokinetics and material balance of diethylenetriamine trihydrochloride ([14C]-DETA·3HCl) in the Fischer 344 rat following oral, endotracheal or intravenous dosing was studied comparabel to an OECD Guideline for Testing of Chemicals No. 417 (2010) study. The bioavailability of [14C]-DETA·3HCl was over 90%. [14C]-DETA·3HCl was well absorbed following oral or endotracheal dosing. Clearance of absorbed [14C]- DETA·3HCl from the plasma was also rapid, with a terminal half-life of 9-16 h. The highest radioactivity concentrations of [14C]- DETA·3HCl was attained in the liver and kidney, suggesting that these organs were common targets for these two alkyleneamines. Metabolism of [14C]-DETA·3HCl appeared to play a small role in the elimination of DETA from the body, as unchanged DETA was the principal component excreted in the urine. The major excretory route of DETA was via the urine and feces. These two pathways together accounted for over 70% of the total dose excreted.

Reference 2

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

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

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Justification for type of information:

The lipolytic activity of human gastric and duodenal juice against medium chain and long chain triglycerides is summarized due to presence of Mono-, di- and triglycerides of fatty acids C16-18 as well as free fatty acids C16-18 within the registered UVCB substance.

Objective of study:

metabolism

Qualifier:

no guideline followed

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

Species:

other: human

Route of administration:

other: in vitro testing

Vehicle:

unchanged (no vehicle)

Conclusions:

The work confirmed extensive literature showing that gastric juice of humans contains lipolytic activity, that ingested mono, di- and triglycerides are 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.
From the data presented it can be expected that > 30% of the mono-, di- and triglycerides of fatty acids C16-18 present in the registered UVCB substance are hydrolyzed by gastric and duodenal juice.

It is considered likely, that the mono-, di- and triglyceride components present in the target substance Amidoamine 2 (UVCB, low nitrogen) will by hydrolzed as described in the this study by lipase by gastric and duodenal juices.

Reference 3

Endpoint:

basic toxicokinetics in vitro / ex vivo

Type of information:

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

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Justification for type of information:

Rearrangement of Glyceride Fatty Acids During Digestion and Absorption is summarized due to presence of Mono-, di- and triglycerides of fatty acids C16-18 as well as free fatty acids C16-18 within the registered UVCB substance.

Objective of study:

absorption

Qualifier:

no guideline followed

Principles of method if other than guideline:

Triglycerides of known structure were fed to rats, and the distribution of the fatty acids in the triglycerides of the lymph was determined.

GLP compliance:

no

Specific details on test material used for the study:

All of the fatty acids used in these studies were analyzed by gas-liquid chromatography and were found to be of better than 99% purity.

The simple triglycerides, triolein and trilinolein, were prepared by transesterification of an excess of the methyl esters of the fatty acids with glycerol in the presence of NaOH. Excess methyl esters were removed by distillation. The asymmetric triglyceride, glyceryl alpha-[palmitate-l-C14] dioleate (palmitate-oleate-oleate), was prepared by acylating alpha-monopalmitin-l-Cl4 that had been prepared by the method of Fischer, Bergmann, and Barwind with oleoyl chloride. The symmetrical triglyceride, glyceryl beta-[palmitate-l-C14] dioleate (oleate-palmitate-oleate) was prepared by acylating beta-monopalmitin-l-Cl4 that had been prepared by the method of Martin with oleoyl chloride. The mixed triglycerides were purified by several recrystallizations.
The iodine value, hydroxyl value, and percentage of free fatty acid were determined on all triglycerides. All these values were within 1% of theory. The proportion of the labeled acid that was in the beta position of the mixed triglycerides was determined by the method of Mattson and Volpenhein . Both triglyceride samples were found to contain no more than 1% of triglycerides of structures other than those intended.

Radiolabelling:

yes

Species:

rat

Strain:

not specified

Details on species / strain selection:

Age: young adults

Sex:

male

Route of administration:

oral: gavage

Vehicle:

not specified

Details on absorption:

The extent of absorption of palmitic acid depended on the form in which it was fed (rates between 52 an 96 %).
Absorption was greatest when palmitic acid was fed as β-palmitoyl diolein, and least when it was fed as the free acid.

Metabolites identified:

yes

Details on metabolites:

The results obtained showed that 85 to 90% of the fatty acids occupy the same position on the lymph triglyceride molecule as they did on the dietary triglyceride molecule before the processes of digestion and absorption.

Rats were fed glyceryl α-[palmitate-1- C14] dioleate, glyceryl β-[palmitate-1- C14] dioleate, a mixture of oleic or linoleic acid and palmitic-1-C14acid, or a mixture of triolein or trilinolein and palmitic-1- C14acid. The position of the palmitic acid on the glycerol in the triglycerides of the lymph was determined.

When the mixtures of free acids were fed, the palmitic acid was essentially randomly distributed among all three positions of the lymph triglyceride molecules. From 85 to 90% of the palmitic acid in the dietary mixed triglycerides was found in its original position on the triglyceride molecule after the processes of digestion and absorption.

When the mixtures of triglyceride and free palmitic acid were fed, 22% of the palmitic acid in the lymph triglycerides was found to be esterified with the β position.

Table 1: Specific activity of lymph triglycerides as percentage of specific activity of fed lipids

Substance fed	%*
Oleic acid (2 moles) and 1 mole of palmitic-1-C14acid	64, 73, 71
Linoleic acid (2 moles) and 1 mole of palmitic-1-C14acid	58, 58, 63
Palmitate-oleate-oleate triglyceride	85, 85, 81
Oleate-palmitate-oleate triglyceride	94, 96, 92
Oleic acid (2 moles) as triolein and 1 mole of palmitic-1-C14acid	59, 54, 54
Linoleic acid (2 moles) as trilinolein and 1 mole of palmitic-1-C14acid	52, 54, 53
*Each value is the percentage obtained in a single experiment

Conclusions:

Absorption rates were between 52 an 96%, depending on the lipid fed. From the data presented it can be expected that > 90% of the mono-, di- and triglycerides of fatty acids C16-18 as well as free fatty acids C16-18 within the registered UVCB substance are absorbed and are available for fat and fatty acid metabolism.

Reference 4

Endpoint:

dermal absorption in vitro / ex vivo

Type of 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:

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

Justification for type of information:

Publication on percutaneous absorption of anionic surfactants (soaps) is presented due to presence in and potential release of fatty acids C16-18 from the registered substance due to metabolic activation and presence of small quantities within the UVCB.

Qualifier:

no guideline followed

Principles of method if other than guideline:

In vitro penetration through rat skin

GLP compliance:

no

Radiolabelling:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

Weight at study initiation: 100-120 g

Type of coverage:

open

Vehicle:

water

Remarks:

The test materials were applied as soap solutions (sodium salts).

Duration of exposure:

24 h

Doses:

Nominal doses: 300 µg C10:0, 325 µg C12:0, 375 µg C14:0, 425 µg C16:0 and 450 µg C18:0/4.9 cm² skin- Dose volume: 0.051 ml/cm²

No. of animals per group:

not specified

Control animals:

no

Details on study design:

Source of skin: rat dorsal skin
Type of skin: full thickness skin
Preparative technique: Dorsal skin was clipped 24 h before cervical dislocation. The skin was excised and mounted in 2.5 cm penetration cells similar to those described by Ainsworth (J Soc Cosmet Chem 11:69 (1960)).
Assay: Diffusion cell, similar to those described by Ainsworth (J Soc Cosmet Chem 11:69 (1960))
Receptor fluid: saline

Signs and symptoms of toxicity:

no effects

Dermal irritation:

no effects

Key result

Time point:

24 h

Dose:

86.73 µg fatty acid C16:0/cm²

Parameter:

percentage

Absorption:

0.2 %

Key result

Time point:

24 h

Dose:

91.84 µg fatty acid C18:0/cm²

Parameter:

percentage

Absorption:

< 0.1 %

The results show no measurable penetration of the C18:0 soap through rat skin up to 24 h after application, but 0.2 µg/cm² of the C16:0 soap had penetrated at 24 h. At the end of the experiment,i .e. 24 h after application, between 60 and 70% of the applied [14C] soaps were rinsed from the skin and 30-40% was associated with the skin.

Conclusions:

The in vitro penetration of C16 and C18 fatty acids (as aqueous sodium salt solutions) through rat skin decreases with increasing chain length. At 86.73 µg fatty acide C16/cm² and 91.84 µg fatty acid C18/cm², about 0.2% and less than 0.1% of the C16 and C18 sodium soap solutions is absoberd after 24 h exposure, respectively. The dermal absorption of fatty acids C16 and C18 present in the substance Triglycerides, C16-18 (even-numbered), reaction products with diethylenetriamine is considered to be low.

Executive summary:

The in vitro penetration of C16 and C18 fatty acids (as aqueous sodium salt solutions) through rat skin decreases with increasing chain length. Based on these findings the dermal absorption of fatty acids C16 and C18 present in the substance Triglycerides, C16-18 (even-numbered), reaction products with diethylenetriamine is considered to be low.

Reference 5

Endpoint:

dermal absorption in vivo

Type of 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:

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

Justification for type of information:

A comprehensive study summary on dermal absorption of Lauramide diethanolamine is presented. The dermal absorption of the test item was assessed based on the physicochemical parameters and information on toxicokinetic literature from structural analogue substances. In summary the substance is anticipated to be orally and dermally absorbed to a moderate to high extent, whereas inhalation is very unlikely.

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 427 (Skin Absorption: In Vivo Method)

Deviations:

not specified

GLP compliance:

not specified

Specific details on test material used for the study:

Target substance:

Amidoamine 2 (UVCB, low nitrogen):
Former chemical name: Amides, from diethylenetriamine and hydrogenated palm oil
CAS No.: 1618093-67-6
New chemical name: Glycerides, C16-18 (even numbered) and their amidation products with diethylenetriamine
Physical state: pale yellowish solid at 20 °C
Batch No.: PU50070067
Purity: 100 % (UVCB)
Storage condition of test material: Room temperature, protected from light
Stability: stable under test conditions

Radiolabelling:

yes

Species:

rat

Strain:

Fischer 344

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Raleigh, NC)
- Age at study initiation: 81 to 87 d
- Housing: Individual glass metabolism chambers, which allowed separate collection of carbon dioxide, urine, and feces.
- Individual metabolism cages: Yes
- Diet: Purina Rodent Chow (no. 5002), ad libitum
- Water: Ad libitum

Type of coverage:

other:

Vehicle:

ethanol

Duration of exposure:

72 h

Doses:

- Actual doses: 25 and 400 mg/kg bw
- Dose volume: 200 and 100 µL of 25 and 400 mg/kg bw respectively

No. of animals per group:

Four

Control animals:

no

Details on study design:

DOSE FORMULATION:
- 16 to 18 µCi radiolabel, an appropriate amount of unlabelled LDEA and 95 % ethanol for a total volume of about 200 µL per dose

VEHICLE:
- Concentration (if solution): 95 %

TEST SITE:
- Preparation of test site: Application site had been clipped of hair the previous day
- Area of exposure: 1 x 1 inch
- Type of cover / wrap if used: A non-occlusive protective appliance, glued over the dose area with cyanoacrylate adhesive.

SITE PROTECTION / USE OF RESTRAINERS FOR PREVENTING INGESTION:
- Not used

REMOVAL OF TEST SUBSTANCE:
- Dose site skin was excised and thoroughly rinsed with ethanol, then gently wiped with cotton gauzes soaked with soapy water.

SAMPLE COLLECTION:
- Gauzes and aliquots of urine, dermal rinse solutions, feces, tissues (liver and kidney) and digested skin samples (in 2N ethanolic sodium hydroxide) were collected and analysed for radiochemical content by adding to vials containing scintillation cocktail (Ultima Gold, Packard Instrument Company).

ANALYSIS
- Method type(s) for identification: Liquid scintillation counting

Details on in vitro test system (if applicable):

not applicable

Signs and symptoms of toxicity:

not examined

Dermal irritation:

not examined

Absorption in different matrices:

- Absorption was relatively slow, and only about 21 to 26 % of the dose penetrated the skin in 72 h and 3 to 5 % remained associated with the dose site.
- No significant differences were observed between doses in the absorption and elimination of LDEA when calculated on a percentage of dose basis.

Total recovery:

- Total recovery: 95 and 92 % recovery was observed at 25 and 400 mg/kg bw respectively

Key result

Time point:

72 h

Dose:

25 mg/kg bw

Absorption:

ca. 26.3 %

Remarks on result:

other: In 25 mg/kg dose group, disposition of radioactivity after dermal application of radiolabelled LDEA to male rats.

TBRs (tissue to blood ratio) were higher in liver (30-40) and kidney (ca.20) than adipose tissue (6-7) and non-dose site skin (ca.2).

Results of jugular vein-cannulated rats, dermally dosed at 100 mg/kg bw of LDEA:

-Only LDEA and the half-acid amide metabolites were detected in plasma, and their levels were near maximal at about 24 h after dosing, with no marked changes in the profiles thereafter.

- Most of the circulating LDEA equivalents were comprised of the two metabolites, with about 15% present as LDEA.

Conclusions:

The study is considered supportive for the hypothesis that based on chemical similarities of source and target substance (tertiary fatty acid amides) the dermal absorption of the test substance in rats is assumed to be low to moderate (<= 30 %). This read-across approach is presented due to the absence of relevant in vivo or in vitro data.
It is considered likely, that the target substance Amidoamine 2 (UVCB, low nitrogen) will show a comparable dermal absorption as the source substance based on the presence of amide and amino functional groups and amphiphilic properties.

Executive summary:

A dermal absorption study was conducted to evaluate the absorption of the radiolabelled (14C) lauramide diethanolamine (LDEA) in the Fischer 344 rats. 25 (200 µL) and 400 mg/kg bw (100 µL) of LDEA were applied to an area of 1x1 inch previously clipped skin, covered with a non-occlusive patch. After 72 h of exposure, gauzes and aliquots of urine, feces, dermal rinse solutions and digested skin samples (in 2N ethanolic sodium hydroxide) were collected and analysed for radiochemical content.

Absorption was relatively slow, and only about 25 to 30% of the dose penetrated the skin in 72 h and 3 to 5% remained associated with the dose site. No significant differences were observed between doses in the absorption and elimination of LDEA when calculated on a percentage of dose basis. The study indicates that the dermal application of the test substance in rats resulted in moderate (25-30%) absorption. Based on a comparable chemical structures (tertiary amides) the dermal absorption of the registered substance monoamide components can be expected.

Description of key information

Introduction

 

Toxicokinetic parameters such as uptake, distribution, metabolism and excretion form part of the essential toxicological profile of a substance. Although the toxicokinetic behaviour of substances does not describe a toxicological endpoint itself, an approximate indication of the individual toxicokinetic parameters absorption, distribution, metabolism and excretion (ADME) can be gained from the results of basic toxicity testing. In this respect the following evaluation discusses results and observations from basic toxicity studies which can be used as approximate indications for the description of every individual toxicokinetic parameter. Such an approach is also justified by animal welfare considerations because additional animal testing can be avoided and is not required in this tonnage band. The assessment of the toxicokinetic properties of the test item given below is based on the results obtained for the toxicological endpoints mentioned below.

There are no studies available in which the toxicokinetic behaviour of the test item has been investigated.

An acute oral toxicity study was performed in rats with the test item accoridng to OECD guideline 423 indicating no markedly signs of systemic toxicity resulting in acute oral LD50 value >2000 mg/kg bw.

In addition, an acute dermal toxicity study in rats and a read across skin sensitisation study was performed with test item according to OECD guideline 402 and 406 demonstrating no signs of systemic toxicity resulting in an acute dermal LD50 value >2000 mg/kg bw and no markedly skin sensitizing reactions.

Moreover, a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test with the test item was performed according to OECD guideline 422 in rats via the oral route at dose levels up to 1000 mg/kg bw/day. No markedly signs of systemic toxicity were observed.

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), assessment of the toxicokinetic behaviour of the test item is conducted to the extent that can be derived from the available information.

A screening summary on oral and dermal absorption of test item is presented. The dermal absorption of the test item was assessed based on the physicochemical parameters and information on toxicokinetic literature from structurally similar substances (lauramide DEA).

In summary the test item is anticipated to be orally absorbed to a high extent (100 %), whereas inhalation uptake is very unlikely due to low vapour pressure and state of matter during application. Moderate dermal absorption (30%) based on experimental data of analogue amides and high lipophilic properties, moleculare weight distribution of components as well as very low water solubility is expected. The substance may be distributed within the organism but accumulation is considered unlikely. Enzymatic hydrolysis by currently unknown protease and enzymase type enzyms is considered likely at the amide or ester site of the test item's components causing it to split in a polar and apolar part. Further phase I and II biotransformation reaction are considered possible.

The absorption of highly lipophilic substances (log P of 4 or above) may be limited by the inability of such substances to dissolve into GI fluids and hence make contact with the mucosal surface. However, the absorption of such substances will be enhanced if they undergo micellular solubilisation by bile salts (Aungst and Shen, 1986). Substances absorbed as micelles (aggregate of surfactant molecules, lowering surface tension) enter the circulation via the lymphatic system, bypassing the liver. Micellular solubilisation is considered a possible path of GI absorbtion of the substances contained in the test item. The major path of excretion is expected to be via bile, although some excretion of polare degradation products via the kidney seems also possible.

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Absorption rate - oral (%):

100

Absorption rate - dermal (%):

30

Absorption rate - inhalation (%):

100