Sodium N-lauroylsarcosinate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04 Aug - 05 Dec 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

adopted 22 January 2001

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Version / remarks:

adopted August 1998

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Japanese Ministry of Agriculture, Forestry and Fisheries Testing guidelines for Toxicology studies, 12 NohSa No 8147, 24 November 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Sodium N-lauroylsarcosinate

EC Number:

205-281-5

EC Name:

Sodium N-lauroylsarcosinate

Cas Number:

137-16-6

Molecular formula:

C15H29NO3.Na

IUPAC Name:

sodium [dodecanoyl(methyl)amino]acetate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crl:CD® (SD)IGS BR

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Margate, UK
- Age at study initiation: not specified in report, animals purchased time-mated
- Weight at study initiation: 197-268 g (females on arrival, prior to GD3)
- Housing: individual in solid-floor propylene cages with stainless steel lids in a single air-conditioned room, bedding with softfood flakes
- Diet: pellet diet (Rodent 2018C Teklad Global Certified Diet, Harlan; Oxon; UK) ad libitum, environmental enrichment provided in the form of wooden chew blocks and cardboard fun tunnels
- Water: ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 50 ± 20
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 04 Aug 2013 (first day of treatment) To: 22 Aug 2013 (final day of necropsy)

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

distilled water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
the test item was prepared at the appropriate concentrations as a solution in distilled water.

VEHICLE
- Concentration in vehicle: 6, 20 and 50 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The test item concentration in the test samples was determined by high performance liquid chromatography (HPLC/UV) using an external standard technique.The formulations investigated during the study were found to comprise test item in the range of 101% to 104% and, thus, the required content limit ± 10% with reference to the nominal content was met. The test item was found to be stable in the formulations when kept for twenty one days in the refrigerator due to results which met the variation limit of 10% from the time-zero mean. Thus, the results indicate the accurate use of the test item and distilled water as vehicle during this study. The formulations were found to be homogeneously prepared, and sufficient formulation stability under storage conditions was approved.

Details on mating procedure:

- Impregnation procedure: purchased timed pregnant

Duration of treatment / exposure:

Gestation Day 5-19

Frequency of treatment:

daily, 7 days/week

Duration of test:

until Gestation Day 20 (terminal sacrifice)

Doses / concentrations

Remarks:

Doses / Concentrations:
30, 100 and 250 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

24 P females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were chosen based on previous toxicity data.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily
- Cage side observations checked included: following arrival, all animals were examined for overt signs of toxicity, ill-health or behavioural changes once daily during the gestation period. An additional observation was also performed five hours after dosing during the normal working week. All observations were recorded.

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: individual body weights were recorded for each surviving individual animal at Day 3, 5, 6, 7, 8, 11, 14, 17 and 20 (termination kill) of gestation.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined: Yes, at Day 3, 5, 8, 11, 14, 17 and 20 of gestation.

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: daily by visual inspection of water bottles
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: ovaries and uteri of pregnant females, full external and internal examination; any macroscopic abnormalities were recorded; the stomach was retained from all females

Ovaries and uterine content:

The ovaries and uterine content were examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

- Other: fetal sex, placental weight, position and type of intrauterine implantation

Fetal examinations:

- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [all per litter]
- Skeletal examinations: Yes: [all per litter]
- Head examinations: No

Statistics:

Group mean values were calculated to include data from all females with live fetuses on Day 20 of gestation. As the litter was the standard unit of assessment, values were first calculated within the litter, and group mean values represent the mean of these individual litter values.
The following parameters were analyzed statistically, where appropriate, using the test methods outlined below:
Female body weight change, food consumption and gravid uterus weight: Bartlett´s test for homogeneity of variance and one way analysis of variance, followed by Dunnett´s multiple comparison test or, if unequal variances were observed, an alternative multiple comparison test.
All caesarean necropsy parameters and fetal parameters: Kruskal-Wallis non-parametric analysis of variance; and a subsequent pairwise analysis of control values against treated values using the Mann-Whitney ‘U’ test, where significance was seen.
Fetal evaluation parameters, including skeletal or visceral findings: Kruskal-Wallis non-parametric analysis of variance and Mann-Whitney ‘U’ test.
Probability values (p) demonstrating different levels of significance were chosen as follows:
p<0.001 ***
p<0.01 **
p<0.05 *
p>=0.05 (not significant)

Indices:

Pre implantation loss: [(number of corpora lutea-number of implantations) / number of corpora lutea] x 100;
Post-implantation loss: [(number of implantations–number of live foetuses)/number of implantations] x 100;
Sex ratio: % male fetuses (sex ratio)= [Number of male foetuses / Total number of foetuses] x 100

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

250 mg/kg bw/day: the female found dead on Day 10 did not show any clinical signs prior to Day 10. The female found dead on Day 18 had noisy respiration on Days 10 and 17 and increased salivation and pilo-erection on Day 17.
5/22 females showed incidences of increased salivation between Days 10 and 19 and 3/22 females also showed noisy respiration on either Day 6 or 7.

100 mg/kg bw/day: 1/24 females showed increased salivation on Day 18 and 1/24 females had noisy respiration on Day 13.

Observations of this nature are commonly observed following the oral administration of an unpalatable or irritant test item formulation and are considered not to represent true systemic toxicity.

Control: 1/24 females had fur loss on Day 18. In the absence of treatment this was considered of no toxicological importance.

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, treatment-related

Description (incidence):

250 mg/kg bw/d: One female treated was found dead on Day 10 and another female was found dead on Day 18.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

250 mg/kg bw/day: body weight gain of the females was lower throughout the treatment period and cumulative body weight gain to Day 14 (where body weight is not greatly influenced by the weight of the gravid uterus) was 24% lower than the corresponding control value. By termination on Day 20, cumulative body weight gain was 15% lower than the corresponding control values. When final body weight was adjusted for the gravid uterus, an overall body weight gain reduction was evident during gestation.

100 mg/kg bw/day: females showed a slight reduction in cumulative body weight gain from Day 7 onwards. When final body weight was adjusted for the gravid uterus, an overall body weight gain reduction was evident during gestation.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

250 mg/kg bw/day: females showed a reduction in food consumption throughout the treatment period. Statistically significant reductions were evident between Days 8 and 11 (p < 0.001) and Days 14-17 (p< 0.05) compared with the control group.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Description (incidence and severity):

The female treated with 250 mg/kg bw/day that was found dead on Day 10 had gaseous distention in the stomach and gastro-intestinal tract. The other interim death female had sloughing on the non-glandular region of the stomach, seven dead foetuses in the right uterine horn and five dead foetuses in the left uterine horn. The remaining females treated with 250 mg/kg bw/day that were terminated on Day 20 all had sloughing on the non-glandular region of the stomach.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Maternal developmental toxicity

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not examined

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not examined

Changes in number of pregnant:

not examined

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

30 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

LOAEL

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

effects observed, treatment-related

Description (incidence and severity):

Females from all treatment groups showed a statistically significant reduction (p<0.05 - p<0.01) in the number of corpora lutea. The intergroup differences did not show a true dose related response and were considered to be incidental and unrelated to treatment due to ovulation and mating occurring prior to the administration of the test item. As a consequence of incidentally lower number of corpora lutea in treated females litter size was statistically significantly reduced (p<0.05) in all treated females. The intergroup differences did not show a true dose related response and in the absence of any effect on post-implantation loss and mean fetal weights it supports the assumption that the intergroup differences in litter size was considered to be incidental and of no toxicological importance.

Changes in postnatal survival:

not specified

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Visceral malformations:

no effects observed

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There was no obvious adverse effect of maternal treatment on litter data as assessed by the mean number of implantations, early and late embryonic/fetal deaths and live fetuses or sex ratio, as assessed by percentage male.
For all dose groups, there were no significant treatment-related trends in the proportion of fetuses (or litters) with evidence of visceral or skeletal anomalies. The types of external, visceral and skeletal anomalies were those commonly observed for this type of study. There were no findings that were considered to represent any known malformations.

Effect levels (fetuses)

Dose descriptor:

NOEL

Effect level:

250 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: No adverse effect on developmental toxicity observed at the highest tested dose level.

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

All treated females showed statistically significant reduction of corpora lutea and in consequence reduced litter sizes. The intergroup differences did not show a true dose related response and were considered to be incidental and unrelated due to ovulation and mating occurring prior to the administration of the test item starting on GD5. The absence of any effect on post-implantation loss and mean fetal weights supports the assumption that these intergroup differences in litter size were considered to be incidental and of no toxicological importance. Foetuses of the 250 mg/kg bw/day group showed a statistically significant increase in the percent of foetuses showing one or more ossified forepaw phalanges. Due to the isolated occurrence of this observation it is not considered to be a true developmental effect and it is therefore of no biological importance.

Table A: Group Mean Litter Data Values (Mean ± SD)

Dose Level (mg/kg bw/day)	Number of Litters	Number of Corpora Lutea	Number of Implants	Number of Embryonic/Fetal Deaths			Number of Live Implants
				Early	Late	Total	Male	Female	Total
0 (control)	23	15.2 ± 1.8	13.9 ± 1.6	0.1 ± 0.3	0.1 ± 0.6	0.3 ± 0.7	6.9 ± 1.8	6.8 ± 1.9	13.7 ± 2.1
30	21	14.3* ± 1.8	13.1 ± 2.0	1.0 ± 2.4	0.1 ± 0.4	1.0 ± 2.5	6.0 ± 2.7	6.1 ± 2.4	12.1* ± 3.4
100	24	13.7* ± 1.9	13.0 ± 1.8	0.6 ± 1.3	0.2 ± 0.7	0.8 ± 1.4	6.5 ± 2.1	5.6 ± 1.7	12.1* ± 2.5
250	22	13.9** ± 1.2	13.2 ± 1.1	0.5 ± 1.4	0.2 ± 0.9	0.7 ± 1.6	6.5 ± 2.0	6.0 ± 1.9	12.5* ± 2.2

p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 not significant

Table A: Group Mean Litter Data Values (continued) (Mean ± SD)

Dose Level (mg/kg bw/day)	Implantation Loss (%)		Male foetuses (%)	Mean Male Fetal Weight (g)	Mean Female Fetal Weight (g)	Mean Fetal Weight (g)	Mean Placental Weight (g)	Litter Weight (g)	Total Placental Weight (g)
	Pre	Post
0 (control)	8.4 ± 7.4	2.3 ± 7.2	50.4 ± 11.3	4.117 ± 0.260	3.935 ± 0.264	4.026 ± 0.257	0.564 ± 0.047	54.883 ± 8.672	7.667 ± 1.141
30	8.0 ± 10.0	8.4 ± 19.4	48.6 ± 15.2	4.152 ± 0.405	3.881 ± 0.372	4.005 ± 0.365	0.578 ± 0.155	48.345 ± 13.761	6.729 ± 1.942
100	5.0 ± 5.2	6.9 ± 12.2	53.7 ± 12.8	4.166 ± 0.339	3.886 ± 0.269	4.036 ± 0.289	0.542 ± 0.060	49.142 ± 11.318	6.540 ± 1.503
250	4.8 ± 5.3	5.5 ± 12.5	52.2 ± 13.7	4.221 ± 0.347	4.033 ± 0.327	4.126 ± 0.320	0.540 ± 0.057	51.237 ± 8.200	6.744 ± 1.369

 

Table B: Group Mean Cumulative Body Weight Change Values (Mean ± SD)

Dose Level (mg/kg bw/day)	Number of Animals	Cumulative Body Weight Change (g) from GD5
		GD6	GD7	GD8	GD11	GD14	GD17	GD20
0 (control)	23	4.0 ± 3.4	7.6 ± 3.7	13.0 ± 5.1	33.6 ± 6.6	51.6 ± 8.1	82.3 ± 10.4	127.8 ± 14.9
30	21	5.1 ± 5.4	7.1 ± 8.2	12.9 ± 10.2	34.0 ± 12.9	53.5 ± 15.5	82.2 ± 19.2	127.0 ± 23.4
100	24	2.3 ± 5.3	6.1 ± 5.5	8.8 ± 7.5	27.6 ± 9.9	44.8 ± 13.8	74.2* ± 12.4	114.3* ± 17.2
250	24/22+	0.5 ± 9.0	3.6 ± 10.1	7.3 ± 11.2	21.7** ± 14.6	39.3* ± 17.8	68.2** ± 20.0	109.1** ± 24.7

+ = n=23 on GD10, n=22 on GD18
p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 not significant

 

Table C: Group Mean Gravid Uterus Weight , Adjusted Body Weight and Body Weight Change Values (Mean ± SD)

Dose Level (mg/kg bw/day)	Number of Animals	Body Weight (g) on Days of Gestation		Body Weight Change (g) during GD5-20	Gravid Uterus Weight (g)	Adjusted Body Weight (g) GD20	Adjusted Body Weight (g) Change GD5-20
		GD5	GD20
0 (control)	23	262.6 ± 20.6	390.4 ± 29.8	127.8 ± 14.9	84.663 ± 12.502	305.8 ± 23.4	43.2 ± 8.2
30	21	261.7 ± 12.9	288.7 ± 27.3	127.0 ± 23.4	75.544 ± 18.733	313.2 ± 28.6	51.5 ± 22.7
100	24	260.6 ± 14.4	374.9 ± 24.8	114.3** ± 17.2	75.893 ± 15.615	299.0 ± 20.7	38.4* ± 14.6
250	24/22+	257.6 ± 15.5	368.1 ± 32.4	109.1** ± 24.7	77.535 ± 11.935	290.6 ± 25.0	31.6** ± 18.7

+ = n=23 on GD10, n=22 on GD18
p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 not significant

 

Table D: Group Mean Food Consumption Values (Mean ± SD)

Dose Level (mg/kg bw/day)	Number of Animals	Food Consumption (g/rat/day) between Days of Gestation
		3-5	5-8	8-11	11-14	14-17	17-20
0 (control)	23	23.0 ± 3.1	25.1 ± 2.5	26.5 ± 2.6	26.1 ± 2.8	25.9 ± 3.0	28.0 ± 2.4
30	21	23.6 ± 3.7	24.5 ± 3.9	27.1 ± 3.7	26.5 ± 3.7	26.0 ± 4.2	28.7 ± 3.8
100	24	22.8 ± 2.9	23.9 ± 2.8	24.9 ± 3.3	24.6 ± 3.7	24.5 ± 3.4	27.5 ± 2.9
250	24/22+#	22.3 ± 3.4	22.4 ± 5.3	21.5*** ± 4.7	23.3 ± 4.3	22.6* ± 4.2	25.4 ± 3.8

+ = n=23 on GD10, n=22 on GD18
# = Food consumption not performed on GD 8 and 11 for female Nos. 85 to 90 due to technician error
p<0.001 ***
p<0.01 **
p<0.05 *
p≥0.05 not significant

In conclusion the oral administration of the test substance to pregnant rats by oral gavage during gestation at dose levels of 30, 100 and 250 mg/kg bw/day resulted in treatment related effects detected in females treated with 250 and 100 mg/kg bw/day. The NOAEL was therefore considered to be 30 mg/kg bw/day.

No toxicological significant changes were detected in the offspring parameters measured. The NOEL for developmental toxicity was therefore considered to be 250 mg/kg bw/day. 

Applicant's summary and conclusion

Conclusions:

Under the conditions of the study the test substance did not induce any treatment-related biologically relevant malformations in the developing unborn organism in the presence of maternal toxicity. Therefore, the test substance does not meet the criteria for classification for prenatal developmental toxicity according to Regulation (EC) No 1272/2008. The available data is thus conclusive but not sufficient for classification.