Oxidised Crude Tall Oil

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route

Endpoint conclusion:

no study available

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

In the available studies no effects on male and female reproductive organs were observed. The substance is of very low toxicity and exposure is expected to be limited.

Effects on developmental toxicity

Description of key information

Female rats (25/dose) were mated and were (after copulation) treated with the test substance at 0, 100 , 300 and 1000 mg/kg bw during day 6 -19 of gestation. No effects on mortality, pregnancy rate, clinical signs, body weight (gain), food consumption and macroscopy were identified. The number of resorptions, implantations, implantation loss and viable fetuses did not differ among groups. Fetal body weights were slightly, but significantly decreased at 1000 mg/kg bw when compared to controls. When compared to historical controls fetal body weights were shown to be within normal ranges. No effects of the substance were found in investigations for external, visceral and skeletal malformation/variations. Therefore it is concluded that the NOAEL for maternal effects is 1000 mg/kg bw. The NOAEL for developmental effects is 1000 mg/kg bw.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 Nov 2014 to 20 Jan 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study under GLP, This Oral (Gavage) Prenatal Developmental Toxicity Study of Tall Oil, Polymd. Oxidized (Envamul™ 200) in Rats was conducted as part of testing plan to meet the China Level III testing requirements.

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

Exposure during day 6-19 of gestation is in agreement with the guidelines. Exposure from fertilization to approximately 1 day prior to the expected day of terminationwas not considered as there are no indications for a high potential for preimplantation loss.

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, NC, USA
- Strain : Crl:CD(SD)
- Age at study initiation: ca. 13 weeks
- Weight at study initiation: 220-280 g
- Fasting period before study: none
- Housing: individually from day 0 of gestation onwards in olid-bottom cage with bedding material
- Diet: PMI Nutrition International, LLC Certified Rodent LabDiet® 5002 ad libitum
- Water: reversed osmosis municipal water ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.3°C to 21.9°C
- Humidity (%): 40.9% to 50.0%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: weekly preparation, stirred continuously throughout the preparation, sampling, and dose administration procedures, stored at room temperature in dark

VEHICLE
- Concentration in vehicle: 0, 10, 30 and 100 mg/mL
- Amount of vehicle (if gavage): 10 mL
- Lot/batch no.: 2DF0279, 2DG0193, and 2DG0376

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Actual concentrations in the test substance formulations were 96.7-99.9% of nominal (samples taken from first and last dose preparation)

System: HPLC-UV
HPLC System: HPLC Agilent 1200 MSD
Column: Phenomenex Luna C8(2), 150 mm × 4.6 mm, 5.0-µm particle-size; Guard Column/Cartridge: Phenomenex Security Guard C18 4.0 × 2.0mm
Column temperature: 40 °C
Detection method: UV (230 nm)
Injection volume: 10 µL
Mobile Phase:Eluate A: 0.1% trifluoroacetic acid in deionized water; Eluate B 0.1% trifluoroacetic acid in acetonitrile
Gradient:
Time(min) A(%) B(%)
0 90 10
2.0 90 10
7.0 10 90
9.0 10 90
9.1 90 10
15.0 90 10
Column flow rate: 1.0 mL/min
Retention time: Approx. 11 min

Calibration solution: 5 mg of the test substance was weighed in a 25-mL flask and brought to volume with ethanol. Aliquots were further diluted with ethanol where necessary
HPLC-UV gave one test substance related peak. No data on the linearity of the system are included.

Analysis for accuracy of preparation, homogenicity and stability:
accuracy of preparation (2 determinations): 96.7-99.9% of nominal
homogenicity (2 determinations): 98.8 ± 2.7% and 97.6 ±0.97% (at 10 mg/mL) and 99.6 ± 1.5% and 101 ±2.7% (at 100 mg/mL)
stability : mean recovery 98-101% over 16 days

Details on mating procedure:

- Impregnation procedure: mated with fertile males before study
- M/F ratio per cage: 1 male/1 female
- Length of cohabitation: max 1 week
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of gestation

Duration of treatment / exposure:

day 6-19 of gestation

Frequency of treatment:

daily

Duration of test:

Day 0 to 20 of gestation

No. of animals per sex per dose:

25 females/dose

Control animals:

yes, concurrent vehicle

Details on study design:

Dose selection rationale: based on findings in an acute oral study (LD50 > 2000 mg/kg bw) and a dose range finding study

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily from day 0 to 20 for mortality and clinical signs (before dosing and 1 hour thereafter)

BODY WEIGHT: Yes
- Time schedule: on day 0 and daily from day 6 to 20

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and daily diet consumption calculated as g food/kg body weight/day: Yes
- Time schedule: on day 0 and daily from day 6 to 20

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Laparohysterectomies and macroscopic examination
- Gravid uterus weight

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number and location of all fetuses, corpora lutea, implantations, early/late resorptions (Salewsky staining where necessary)
- Other: placentae

Fetal examinations:

- Sex (ratio)
- External examinations: all fetuses including sex, eyes, palate, and external orifices; late resorptions crown-rump measurements and degrees of autolysis
- Soft tissue examinations: all fetuses (1/2 of the heads) includes fetal kidney examination for development of papillae
- Skeletal examinations: 1/2 of the fetuses
- Heads: 1/2 of the fetuses midcoronal slice taken

Statistics:

internal system including ANOVA, Dennet's test, Kruskal-Wallis nonparametric ANOVA, Dunn's test,

Indices:

all intrauterine findings were indicated on group mean litter basis and proportional litter basis
all fetal results were indicated as incidence per group as well as as affected fetuses per litter

Historical control data:

included in an appendix of the report

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Increased incidences of soft feces were noted in
the 300 and 1000 mg/kg/day groups compared to the control group sporadically
throughout the treatment period (gestation days 7-20) at the daily examinations; however,
given the low occurrence of this finding (generally 1-4 occurrences per animal), it was
considered test substance-related but nonadverse. In the 1000 mg/kg/day group, all
females were noted with brown and/or red material around the mouth during gestation
days 7-19 at approximately 1 hour following dose administration. However, these
findings did not persist to the daily examinations. Furthermore, the brown material
findings correlated with the color of the test substance (dark brown, opaque liquid).
Therefore, the brown and/or red material findings were considered test substance-related
but nonadverse. No other test substance-related clinical findings were noted at the daily
examinations or approximately 1 hour following dose administration at any dosage level.
Findings noted in the test substance-treated groups, including hair loss on various body
surfaces and yellow material on the urogenital and anogenital areas, occurred
infrequently, at similar frequencies in the control group, and/or in a manner that was not
dose-related.
see attached tables

Mortality:

no mortality observed

Description (incidence):

All females in the control, 100, 300, and 1000 mg/kg/day groups survived to the scheduled necropsy on gestation day 20.
see attached tables

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Significantly (p<0.05 or p<0.01) higher mean body weight gains were noted in the 100 mg/kg/day
group on gestation day 6-7 and in the 300 mg/kg/day group on gestation day 18-19
compared to the control group; these differences were transient, did not occur in a
dose-related manner, and were not of sufficient magnitude to affect mean body weights in
these groups. Therefore, these higher mean body weight gains were not considered test
substance-related.
see attached tables

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Maternal food consumption, evaluated as g/animal/day and g/kg/day, in the 100, 300, and
1000 mg/kg/day groups was unaffected by test substance administration. The only
significant (p<0.05 or p<0.01) differences from the control group were higher mean food
consumption noted during gestation day 7-8 at 300 mg/kg/day, gestation day 12-13 at
100 and 1000 mg/kg/day, and gestation days 12-15 at 1000 mg/kg/day (g/kg/day value
only). The aforementioned differences were transient, did not occur in a dose-related
manner, and/or were not of sufficient magnitude to affect mean g/animal/day food
consumption and/or mean body weights in these groups. Therefore, the higher food
consumption in these groups was not considered test substance-related.
see attached table

Food efficiency:

not examined

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

not examined

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:

no effects observed

Description (incidence and severity):

No effects on gravid uterus weight were observed
see attached tables

Gross pathological findings:

no effects observed

Description (incidence and severity):

At the scheduled necropsy on gestation day 20, no test substance-related internal findings
were observed at dosage levels of 100, 300, and 1000 mg/kg/day. Macroscopic findings
observed in the test substance-treated groups occurred infrequently, at similar frequencies
in the control group, and/or in a manner that was not dose-related. One female each in
the control, 100, and 1000 mg/kg/day groups was determined to be nongravid.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Number of abortions:

no effects observed

Description (incidence and severity):

no abortions were observed, one female in the control, 100, and 1000 mg/kg/day groups was determined to be nongravid.

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

Intrauterine growth at 100 and 300 mg/kg/day and survival at 100, 300, and
1000 mg/kg/day were unaffected by test substance administration. Parameters evaluated
included postimplantation loss, live litter size, mean fetal body weights, and fetal sex
ratios. Mean numbers of corpora lutea and implantation sites and the mean litter
proportions of pre-implantation loss were similar across all groups. Differences from the
control group were slight and not statistically significant.
see attached tables

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

Intrauterine growth at 100 and 300 mg/kg/day and survival at 100, 300, and
1000 mg/kg/day were unaffected by test substance administration. Parameters evaluated
included postimplantation loss, live litter size, mean fetal body weights, and fetal sex
ratios. Mean numbers of corpora lutea and implantation sites and the mean litter
proportions of pre-implantation loss were similar across all groups. Differences from the
control group were slight and not statistically significant.
see attached tables

Early or late resorptions:

no effects observed

Description (incidence and severity):

Intrauterine growth at 100 and 300 mg/kg/day and survival at 100, 300, and
1000 mg/kg/day were unaffected by test substance administration. Parameters evaluated
included postimplantation loss, live litter size, mean fetal body weights, and fetal sex
ratios. Mean numbers of corpora lutea and implantation sites and the mean litter
proportions of pre-implantation loss were similar across all groups. Differences from the
control group were slight and not statistically significant.
see attached tables

Dead fetuses:

no effects observed

Description (incidence and severity):

The numbers of fetuses (litters) available for morphological evaluation were 345(23), 367(24), 376(25), and 347(24) in the control, 100, 300, and 1000 mg/kg/day groups, respectively.
see attached tables

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:

no effects observed

Description (incidence and severity):

One female in the control, 100, and 1000 mg/kg/day groups was determined to be nongravid.

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
Mortality: none
Clinical signs: no treatment related effects (incidentally soft stool at 300 and 1000 mg/kg bw and red discoloration of the snout at 1000 mg/kg bw)
Body weight (gain): no treatment related effects
Food consumption: no treatment related effects
Pregnancy: 24, 24, 25 and 24 at 0, 100, 300 and 1000 mg/kg bw
Viable fetuses: no treatment related effects
Pre-implantation loss: no treatment related effects
Early/late resorptions: no treatment related effects
Post implantation loss: no treatment related effects
Fetal body weights: significantly decreased at 1000 mg/kg bw (males and females, but within historical control ranges)
Placentae: no treatment related effects

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Abnormalities:

no effects observed

Fetal body weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

In the 1000 mg/kg/day group, mean male, female, and combined fetal weights (3.6 g,
3.5 g, and 3.6 g, respectively) were 7.7%, 5.4%, and 5.3% lower, respectively, compared
to the control group (3.9 g, 3.7 g, and 3.8 g, respectively); differences were significant
(p<0.05 or p<0.01). However, this effect on fetal weights at 1000 mg/kg/day did not
result in a test substance-related effect on fetal morphology at this dosage level and were within historical control ranges
see attached tables
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

Description (incidence and severity):

live off spring: 345, 367, 376 and 347 (91.6, 96.6, 95.0 and 93.9%/litter) at 0, 100, 300 and 1000 mg/kg bw
see attached table

Changes in sex ratio:

no effects observed

Description (incidence and severity):

Male/female 54/46, 51/49, 46/54, and 55/45 at 0, 100, 300 and 1000 mg/kg bw
see attached tables

Changes in litter size and weights:

not examined

Changes in postnatal survival:

not examined

External malformations:

no effects observed

Description (incidence and severity):

There were no external malformations or developmental variations. The numbers of fetuses (litters) available for morphological evaluation were 345(23),
367(24), 376(25), and 347(24) in the control, 100, 300, and 1000 mg/kg/day groups, respectively
see attached table

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There were no test substance-related skeletal malformations noted for fetuses in the test
substance-treated groups. In the 300 mg/kg/day group, 1 fetus was noted
with severely malaligned sternebrae. Because this malformation was noted for a single
fetus, did not occur in a dose-related manner, and the mean litter proportion was within historical
control data range, it was not considered test substance-related.
No test substance-related skeletal developmental variations were noted for fetuses in the
100, 300, and 1000 mg/kg/day groups. Skeletal developmental variations that were noted
in all groups, including the control group, consisted of cervical centrum no. 1 ossified,
14th rudimentary rib(s), sternebra(e) nos. 5 and/or 6 unossified, hyoid unossified, and
sternebra(e) malaligned (slight or moderate). These findings were noted similarly in the
concurrent control group, did not occur in a dose-related manner, and/or the mean litter
proportions were within the WIL Research historical control data ranges. Other skeletal
developmental variations observed in the test substance-treated groups were noted in
single fetuses, occurred infrequently, did not occur in a dose-related manner, and/or the
mean litter proportions were not statistically significantly different from the concurrent
control group and were within the WIL Research historical control data ranges.
Therefore, these findings were not considered test substance-related.
see attached tables

Visceral malformations:

no effects observed

Description (incidence and severity):

No visceral malformations were noted for fetuses at any dosage level.
No test substance-related visceral developmental variations were noted in the 100, 300,
and 1000 mg/kg/day groups. Renal papilla(e) not fully developed and/or distended
ureter(s) were noted for 1, 2, and 6 fetuses in the control, 300, and 1000 mg/kg/day
groups, respectively. Two and 1 fetuses in the 100 and 1000 mg/kg/day groups,
respectively, were noted with major blood vessel variation (right subclavian and right
carotid arteries arose independently from the aortic arch [no brachiocephalic trunk]). In
the 300 mg/kg/day group, a single fetus was noted with a small and pale spleen. Visceral
findings in the test substance-treated groups occurred infrequently or at a frequency
similar to the concurrent control group, did not occur in a dose-related manner, and/or the
mean litter proportions were within the WIL Research historical control data ranges;
therefore, they were not considered test substance-related.
In the 1000 mg/kg/day group, 4 fetuses were noted with renal papilla(e) not fully
developed (Woo and Hoar Grade 1) and 1 fetus was noted with a cystic urinary bladder.
These findings were not classified as either a malformation or developmental variation,
were not included on the summary tables, and were not considered to be test
substance-related because they occurred infrequently.
see attached tables

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Viable fetuses: 345, 367, 376 and 347 (91.6, 96.6, 95.0 and 93.9%/litter) at 0, 100, 300 and 1000 mg/kg bw
Fetal body weights: significantly decreased at 1000 mg/kg bw (males and females, but within historical control ranges)
Malformations: no external and no visceral malformations, 1 fetus at 300 mg/kg bw with malaligned sternebrae (skeletal malformation)
Variations: incidental with no relationship with treatment and/or at frequencies similar as observed in controls

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Maternal data

Dose (mg/kg bw)	0	100	300	1000	Treatment related
Endpoint
Mortality	No mortality				No
Clinical signs - hair loss forelimbs - soft faeces - red material around nose	7 2 1	10 4 1	7 8 2	9 8 2	No (incidental findings)
Pregnancy rate	24/25	24/25	25/25	24/25	No
Abortions	0	0	0	0	No
Body weight gain d 6-20 (g)	107	113	112	107	No
Body weight gain d 1-20 corr. for gravid uterus weight (g)	60.0	61.5	62.2	59.0	No
Gravid uterus weight (g)	83.7	87.4	87.4	81.5	No
Food consumption	NTRE				No
Macroscopy	NTRE				No
Corpora Lutea	16.2	16.7	17.0	16.5	No
Implantation sites	15.1	15.8	15.8	15.4	No
Early resorptions (%)	8.2	3.4	5.0	6.1	No
Late resorptions (%)	0.2	0	0	0	No
Pre-Implantation loss (mean %)	8.0	4.6	6.8	6.5	No
Post-Implantation loss (mean %)	8.4	3.4	5.0	6.1	No
Viable foetuses (%/mean)	91.6/14.4	96.6/15.3	95.0/15.0	93.9/14.5	No
Dead foetuses (%)	0	0	0	0	No

NTRE= no treatment related effects

↓statistically decreased

Fetal adata

Dose	0		100		300		1000		Treatment related
Endpoint	M	F	M	F	M	F	M	F
Fetal weight (mean g)	3.8		3.7		3.8		↓3.6 (M+F)(-5.3%)		No (within historical control)
Sex ratio (%)	54	46	51	49	46	54	55	45	No
External findings(% litters affected	0		0		0		0		No
Visceral malformations(no foetuses/no litters)	0/0		0/0		0/0		0/0		No
Variations (%/litter)*:	0.9		0.6		0.9		2.0		No, incidental
Skeletal malformations(no foetuses/no litters)	0/0		0/0		1/1		0/0		No
Malformations (no foetuses) - sternebrae malaligned (severe)	0		0		1		0		No
Variations (%/litter)	38.8		35.1		47.2		45.3		No, no relationship with dose

↓statistically decreased

*LIVER- ACCESSORY LOBULE(S),  RENAL PAPILLA(E) NOT DEVELOPED AND/OR DISTENDED URETER(S),  MAJOR BLOOD VESSEL VARIATION, SPLEEN- SMALL, SPLEEN- PALE

** in majority of the litter:sCERVICAL CENTRUM #1 OSSIFIED, 14TH RUDIMENTARY RIB(S), STERNEBRA(E) #5 AND/OR #6 UNOSSIFIED

Incidentally: BENT RIB(S), HYOID UNOSSIFIED, 14TH FULL RIB(S), STERNEBRA(E) #1,#2,#3 AND/OR #4 UNOSSIFIED, STERNEBRA(E) MALALIGNED(SLIGHT OR MODERATE), REDUCED OSSIFICATION OF THE 13TH RIB(S), 27 PRESACRAL VERTEBRAE, REDUCED OSSIFICATION OF THE VERTEBRAL ARCHES, PUBIS UNOSSIFIED, 7TH CERVICAL RIB(S)

Discussion

Based on the lack of adverse maternal effects at any dosage level, a dosage level of

1000 mg/kg/day (the highest dosage level tested) was considered to be the

no-observed-adverse-effect level (NOAEL) for maternal toxicity when EnvaMul™ 200

was administered orally by gavage to bred Crl:CD(SD) rats.  Lower mean fetal weights

were noted in the 1000 mg/kg/day group.  Intrauterine growth at 100 and 300 mg/kg/day

and intrauterine survival, and external, visceral, and skeletal fetal morphology at 100,

300, and 1000 mg/kg/day were unaffected by test substance administration.  Based on

these results, a dosage level of 300 mg/kg/day was considered to be the NOAEL for

embryo/fetal development.

 

Conclusions:

The NOAEL for maternal toxicity is 1000 mg/kg bw
No developmental effects were observed and the NOAEL for developmental toxicity is 1000 mg/kg bw.

Executive summary:

Female rats (25/dose) were mated and were (after copulation) treated with the test substance at 0, 100 , 300 and 1000 mg/kg bw during day 6 -19 of gestation. No effects on mortality, pregnancy rate, clinical signs, body weight (gain), food consumption and macroscopy were identified. The number of resorptions, implantations, implantation loss and viable fetuses did not differ among groups. Fetal body weights were slightly, but significantly decreased at 1000 mg/kg bw when compared to controls. When compared to historical controls fetal body weights were shown to be within normal ranges. No effects of the substance were found in investigations for external, visceral and skeletal malformation/variations. Therefore it is concluded that the NOAEL for maternal effects is 1000 mg/kg bw. The NOAEL for developmental effects is 1000 mg/kg bw.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Justification for selection of Effect on developmental toxicity: via oral route:
Guideline study under GLP

Justification for classification or non-classification

The available data on developmental toxicity and repeated dose toxicity do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.