4-(1,1,3,3-tetramethylbutyl)phenol

Specific investigations: other studies

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

endocrine system modulation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: meets generally accepted scientific standards, well documented and acceptable for assessment

Qualifier:

according to guideline

Guideline:

other: Hershberger Assay (OECD 2001)

Principles of method if other than guideline:

Chemicals are administered orally for 10 days to castrated rats.

GLP compliance:

yes

Type of method:

in vivo

Endpoint addressed:

not applicable

Species:

rat

Strain:

other: Brl Han: WIST Jcl (GALAS)

Sex:

male

Route of administration:

oral: gavage

Vehicle:

olive oil

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

10 d

Frequency of treatment:

daily

Post exposure period:

24-h after final dose

Remarks:

Doses / Concentrations:
50 mg/kg
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
200 mg/kg
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
400 mg/kg
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
600 mg/kg
Basis:
nominal conc.

No. of animals per sex per dose:

6

Control animals:

yes, concurrent vehicle

Hershberger Assay - adminstration of 4-tert-octylphenol with and without testosterone propionate (TP) for 10 days 

Doses (mg/kg per day)	Body Weight (g)	Ventral Prostate (mg/100 g bw)	Seminal vesicle (mg/100 g bw)	BC/LA (mg/100 g bw)	Glans penis (mg/100 g bw)	Cowper’s gland (mg/100 g bw)
Vehicle control	281.9±7.9	5.2±0.7	10.2±1.2	50.0±5.4	11.6±1.3	1.2±0.4
50	278.7±9.5	5.9±0.9	11.6±0.7*	50.8±6.7	12.8±1.4	2.1±1.1
200	256.7±15.3**	5.8±0.9	10.4±1.9	49.4±7.8	12.3±2.1	1.7±0.4
600(400)#	-	-	-	-	-	-
Vehicle control +TP	285.0±14.4	36.4±4.6	76.9±18.3	113.5±12.9	24.0±2.6	6.8±0.9
50+TP	281.5±13.0	36.2±6.2	83.5±13.8	106.9±9.3	23.2±0.8	6.2±1.2
200+TP	277.6±18.7	42.9±5.7	93.3±19.5	119.6±6.2	24.9±1.0	8.2±1.1*
600(400)# +TP	-	-	-	-	-	-
Flutamide +TP	283.1±7.5	6.7±0.9**	10.8±1.7**	50.2±4.2**	11.3±2.6**	1.3±0.3**

#       Numbers in parenthesis are reduced dose because toxic signs were observed during the study.

-          No data because animals died during the study 

*         Significantly different from vehicle control or vehicle plus TP at P<0.05

**        Significantly different from vehicle control or vehicle plus TP at P<0.01

TP      Testosterone propionate (TP, CAS No 57-63-6, 98% purity, Sigma) 0.2 mg/kg per day administered subcutaneous injection into the back after oral administration of the chemical

Conclusions:

In this Hershberger assay study male castrated rats were administered 50, 200 and 600 mg/kg bw octylphenol. Animals receiving 200 mg/kg bw showed decreased body weight gain and decreased spontaneous locomotion. Death occurred at the highest concentration. No significant dose related changes of the sex organ weights were noted. Co-administration with Testosterone propionate caused an increase in the relative weight of the Cowper’s gland at 200 mg/kg day.

Executive summary:

In a study to assess estrogenic and androgenic properties 4-tert-octylphenol was administered to 6 male Brl Han: WIST Jcl (GALAS) rats by oral gavage at dose levels of 50, 200 and 600 mg/kg bw/day. Parallel groups of animals were co-administered with Testosterone propionate (TP).

All rats given 600 mg/kg bw 4-tert-octylphenol as well as those given 600 mg/kg bw plus TP died during the administration period. Treatment with 200 mg/kg bw 4-Tert-octylphenol resulted in significant body weight loss. For all groups, the accessory sex organ weight values are within the control ranges. The relative seminal vesicle weights increased significantly in the low-dose group only (50 mg/kg bw 4-tert-ocylphenol). The relative Cowper’s gland weight increased in rats co-administered with TP at 200 mg/kg bw but an apparent dose dependency was not observed. The seminal vesicle weight in this group was within the control ranges of other studies. From the data provided it cannot be concluded that octylphenol acts as an estrogen agonist or androgen antagonist.

Reference 2

Endpoint:

endocrine system modulation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: meets generally accepted scientific standards, well documented and acceptable for assessment

Reason / purpose for cross-reference:

reference to same study

Qualifier:

no guideline available

Principles of method if other than guideline:

Yeast assay (Saccharomyces cerevisiae) - the oestrogenic activity was expressed as a potency relative to 17ß-estradiol

GLP compliance:

not specified

Type of method:

in vitro

Endpoint addressed:

not applicable

Species:

other: yeast

Strain:

other: Sacccharomyces cerevisiae

Sex:

not specified

Route of administration:

other: assay

Vehicle:

ethanol

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

84 h

Frequency of treatment:

single

Post exposure period:

7 d

Remarks:

Doses / Concentrations:
5-60 mM
Basis:
other: stock solutions of the chemicals were made up in ethanol and diluted to achieve final concentrations

No. of animals per sex per dose:

Duplicates applied into wells of optically flat 96-well microtiter plates.

Control animals:

yes, concurrent vehicle

Details on results:

In a comparative study of alkyl phenols with the main natural estrogen 17ß-estradiol, 4-tert-octylphenol was found to be about 1 000 times less potent than 17ß-oestradiol.

Tamoxifen, an oestrogen antagonist known to act via the estrogen receptor, was shown to inhibit the activity of the alkyl phenols, demonstrating that the assay response was due to interaction with the estrogen receptor.

Conclusions:

In a comparative study of alkyl phenols with the main natural estrogen 17ß-oestradiol, 4-tert-octylphenol was found to be about 1000 times less potent than 17ß-oestradiol.

Executive summary:

In a study on endocrine system modulation, 4 -tert-Octylphenol (99 % a.i.) was tested applied at concentrations of 5 -60 mM to optically flat 96-well microtiter plates containing recombinant yeast as assay medium. After 84 hour incubation, absorbance of 540 nm was read and compared to that of 17ß-estradiol. Control animals received the concurrent vehicle. After the post exposure period of 7 days, 17ß-estradiol was found to be about 1000 times more potent than 4-tert-octylphenol. Tamoxifen, an estrogen antagonist known to act via the estrogen receptor, was shown to inhibit the activity of the alkyl phenols, demonstrating that the assay response was due to interaction with the estrogen receptor.

This study is classified acceptable, it meets generally accepted scientific standards, is well documented and acceptable for assessment.

Reference 3

Endpoint:

endocrine system modulation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Meets generally accepted scientific standards, well documented and acceptable for assessment

Qualifier:

no guideline available

Principles of method if other than guideline:

rat uterine cytosolic (RUC) ER-competitive binding assay

GLP compliance:

not specified

Type of method:

in vitro

Endpoint addressed:

not applicable

Details on results:

This study was conducted to characterize the estrogen receptor (ER)–binding affinity of 4-(1,1,3,3,-Tetramethylbutyl)-phenol using the rat uterine cytosolic (RUC) ER-competitive binding assay, with secondary analysis using Lineweaver-Burk plots and slope replots to confirm true competitive inhibition and to determine an experimental Ki. The inhibitory concentration (IC 50) of 4-(1,1,3,3,-Tetramethylbutyl)-phenol was reported as 12.0 µM compared to 0.00052 µM for 17 -b–Estradiol.

Conclusions:

The inhibitory concentration (IC 50) of 4-(1,1,3,3,-Tetramethylbutyl)-phenol was reported as 12.0 µM compared to 0.00052 µM for 17 -b–Estradiol.

Executive summary:

This study was conducted to characterize the estrogen receptor (ER)–binding affinity of 4-(1,1,3,3,-Tetramethylbutyl)-phenol using the rat uterine cytosolic (RUC) ER-competitive binding assay, with secondary analysis using Lineweaver-Burk plots and slope replots to confirm true competitive inhibition and to determine an experimental Ki. The inhibitory concentration (IC 50) of 4-(1,1,3,3,-Tetramethylbutyl)-phenol was reported as 12.0 µM compared to 0.00052 µM for 17 -b–Estradiol.

This study is acceptable, it meets generally accepted scientific standards, is well documented and acceptable for assessment.

Reference 4

Endpoint:

specific investigations: other studies

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

March 1996

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Meets generally accepted scientific standards, well documented and acceptable for assessment.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Assessment of effects on uterine growth on immature female rats. All animals received a single dose (10 mg/kg bw) Octylphenol via oral gavage on each day for three days. The animals were terminated 24 hours after the final dose. A preliminary range-finding study was conducted to evaluate the maximum tolerated dose.

GLP compliance:

no

Type of method:

in vivo

Endpoint addressed:

toxicity to reproduction / fertility

Species:

rat

Strain:

other: Alpk:APfSD (Wistar-derived)

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:BABU, Alderely Park, Cheshire, UK
- Age at study initiation: (3) x wks; (F1) x wks
- Weight at study initiation: (P) Females: 37-48 g; (F1) Males: x-x g; Females: x-x g
- Housing: 6 per cage
- Diet (e.g. ad libitum): PCD, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 24 h

ENVIRONMENTAL CONDITIONS
- Temperature (°C): standard conditions
- Humidity (%): standard conditions
- Photoperiod (hrs dark / hrs light): standard conditions

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
All dosings were prepared on the day dosing commenced and were stored at room temperature throughout the dosing period.

DIET PREPARATION
- Rate of preparation of diet (frequency): daily
- Mixing appropriate amounts with (Type of food): PCD diet
- Storage temperature of food: room temperature

VEHICLE
- Justification for use and choice of vehicle (if other than water): no data
- Concentration in vehicle: not analyzed
- Amount of vehicle (if gavage): 10 ml/kg

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

3 days

Frequency of treatment:

daily

Remarks:

Doses / Concentrations:
100, 200, 300, 400 mg/kg/bw
Basis:
actual ingested

No. of animals per sex per dose:

6 animals per dose (all females)

Control animals:

yes, concurrent vehicle

other: yes, positiv control: oestradiol benzoate (0.5 µg/animal, subcutaneously)

Details on study design:

The body weight of each animal was recorded upon delivery immediately prior to dosing on each day to termination.
Clinical observations were conducted prior to the start of the study as well as at the time of weighing and dosing. The rats were observed at least once daily (post-dosing) during the study.
Day of termination: 24 h after the third and final dose. An overdose of inhalation anaesthetic (fluorethane) was used. The uterus was removed from each animal, trimmed and blotted on filter paper to remove any fluid. The uterus was then weighed and discarded.

Effect levels

Endpoint	Generation	Sex	Effect level	Based on	Basis for effect level / Remarks
NOAEL	other: immature rats	female	400 mg/kg bw/day (actual dose received)	test mat.	uterine weight (absolute)

There were no significant differences in the group mean terminal body weights except for the two highest doses (300 and 400 mg/kg) which were reduced to 90% and 77% of the vehicle control group, respectively. The group body weight gain was significantly reduced in the 300 mg/kg dose group (63%) while the 400 mg/kg dose group exhibited no measurable body weight gain.

No treatment related effects on clinical signs were observed for any of the dose groups.

No significant effects were observed in absolute uterine weight. The relative uterine:body weight ratio was increased 1.18 -fold in animals treated with 100, 200, and 300 mg/kg and was elevated 1.31 -fold in animals receiving the top dose (400 mg/kg). The small increases in uterine weight in the absence of any increase in absolute uterine weight are considered to be of no biological significance.

Conclusions:

Octylphenol produced a slight but statistically significant increase in relative (not absolute) uterine weight at all dose levels. However, the biological importance of this slight response remains unclear. The results indicate that octylphenol does not possess the intrinsic potential to mimic oestrogen action, under the conditions of this study.

Executive summary:

Octylphenol was administered to 6 immature female Alpk:APfSD (Wistar-derived) rats/dose by gavage at dose levels of 0, 100, 200, 300, and 400 mg/kg bw/day for three consecutive days. A slight but statistically significant increase in relative uterine weight at all dose levels was reported (1.18 - 1.31 -fold). The biological importance of this slight response remains unclear. It cannot be concluded that octylphenol possesses an intrinsic potential to mimic oestrogen action, under the conditions of this study.

The NOAEL on uterine weight is 400 mg/kg bw/day in immature females.

This uterotrophic assay in the rat is classified acceptable for assessment of the estrogenic activity of Octylphenol.

Description of key information

The relative estrogenic potency of Octylphenol (PTOP) was reported in in vitro studies to be 10^3 – 10^5 fold lower compared to estradiol (Routledger & Sumpter, 1997; Laws, 2006). 
From the data of two uterotrophic assays it cannot be concluded that octylphenol acts as estrogen agonist or androgen antagonist (Yamasaki, 2003; ICI Surfactant, 1996). 

Additional information

Estrogen-like effect:

The relative estrogenic potency of Octylphenol (PTOP) was reported in in vitro studies to be 10^3 – 10^5 fold lower compared to estradiol (Routledger & Sumpter, 1997; Laws, 2006). Receptor binding alone is not sufficient to postulate any estrogenic effect.

Two in vivo uterotrophic assays were conducted. ICI Surfactant (1996) reported a small but significant increases in relative (but not absolute) uterine weight at all dose levels (100-400 mg/kg bw). These changes were within historical control values and considered to be of no biological significance. Yamasaki (2003) conducted a similar test at dose levels of 100, 200 and 600 mg/kg bw. All animals of the high dose group died.Treatment with 200 mg/kg bw resulted in significant body weight loss. The accessory sex organ weight values were within the control ranges in all dose groups. The relative seminal vesicle weights increased significantly but not dose-related in the low-dose group only (50 mg/kg bw 4-tert-ocylphenol). The relative Cowper’s gland weight increased in rats co-administered with TP at 200 mg/kg bw but an apparent dose dependency was not observed. The seminal vesicle weight in this group was within the control ranges of other studies. From the data provided it cannot be concluded that octylphenol acts as an estrogen agonist or androgen antagonist.