Reaction mass of 4,4'-[2,2,2-trifluoro-1-(trifluoromethyl)ethylidene]diphenol and benzyltriphenylphosphonium, salt with 4,4'-[2,2,2-trifluoro-1-(trifluoromethyl) ethylidene]bis[phenol] (1:1)

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Determination of bisphenol AF (BPAF) in tissues, serum, urine and feces of orally dosed rats by ultra-high-pressure liquid chromatography–electrospray tandem mass spectrometry

Author:

Yang et al.

Year:

2012

Bibliographic source:

Journal of Chromatography B, 901 (2012) 93– 97

Report date:

2012

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Deviations:

yes

Remarks:

Only 1 dose rate tested with no justification provided as to why this would be sufficient. Presence of metabolites not mentioned.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

4,4'-[2,2,2-trifluoro-1-(trifluoromethyl)ethylidene]diphenol

Cas Number:

1478-61-1

Molecular formula:

C15H10F6O2

IUPAC Name:

4,4'-[2,2,2-trifluoro-1-(trifluoromethyl)ethylidene]diphenol

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Stock solution (in methanol) stored at -20 ºC.
- Stability under test conditions: Not required (ADME study- with UPLC/MS analysis throughout testing period)
- Solubility and stability of the test substance in the solvent/vehicle: Stock solution prepared at 1 mg/L.
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: Not reported

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Dissolved in methanol.
- Preliminary purification step (if any): n/a
- Final dilution of a dissolved solid, stock liquid or gel: 1 mg/L stock solution prepared, in methanol.
- Final preparation of a solid: n/a

FORM AS APPLIED IN THE TEST (if different from that of starting material): Applied as a liquid.

OTHER SPECIFICS: n/a

Radiolabelling:

no

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Academy of Military Medical Sciences, China.
- Age at study initiation: 8 - 9 weeks old
- Weight at study initiation: 250 - 300 g
- Housing: Housed individually in steel metabolism cages.
- Diet (e.g. ad libitum): Ad libitum
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 25 ºC
- Humidity (%): Not reported
- Air changes (per hr): Not reported
- Photoperiod (hrs dark / hrs light): 12:12 light: dark

IN-LIFE DATES: Not reported

Administration / exposure

Route of administration:

oral: unspecified

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Dosing solutions were prepared from a 1 mg/L methanol stock solution. Details on the preparation of the dose solutions are not reported.

DIET PREPARATION
- Rate of preparation of diet (frequency): Not reported
- Mixing appropriate amounts with (Type of food): n/a
- Storage temperature of food: n/a

Duration and frequency of treatment / exposure:

Individuals were treated daily for 2 weeks.

Doses / concentrations

Dose / conc.:

10 mg/kg bw/day (nominal)

No. of animals per sex per dose / concentration:

4

Control animals:

yes

Positive control reference chemical:

No

Details on study design:

- Dose selection rationale: Approximately 350-fold lower than the published acute LD50 data in rats.
- Rationale for animal assignment (if not random): Not reported

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, kidney, liver, testis, adipose and muscle.
- Time and frequency of sampling: At the end of the treatment period.
- Other: n/a
METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled (delete / add / specify): urine, faeces, tissues (as above).
- Time and frequency of sampling: 1 sampling interval at the end of the treatment period (Day 14).
- From how many animals: 4
- Method type(s) for identification HPLC-MS-MS
- Limits of detection and quantification: Signal: noise ratio of 3:1 and 10:1 for LOD and LOQ, respectively, yielded quantification levels of 1 µg/kg for liver, muscle, adipose, kidney and urine samples; and 3 µg/kg for faeces.
- Other: n/a
TREATMENT FOR CLEAVAGE OF CONJUGATES (if applicable): Enzymatic hydrolysis

Statistics:

A statistical analysis was performed by SPSS 16.0 to compare the difference in BPAF concentrations between samples with and without enzymatic hydrolysis, using the Wilcoxon Signed Ranks Test. The results of the analysis would dictate if the enzymatic hydrolysis procedure should be adopted in the test.

Results and discussion

Preliminary studies:

Recovery studies were performed at three fortification levels, which were selected according to the sensitivity of each matrix. The mean recoveries ranged from 71.0% to 102.3% with relative standard deviations of no more than 13.2% (n = 6).

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Bisphenol-AF was poorly adsorbed with the vast majority of the test item excreted in the faeces ( > 10 mg/kg).

Details on distribution in tissues:

Bisphenol-AF was found in highest concentrations in liver and kidney tissues and serum. Maximum concentrations of 1637.5 µg/kg were observed in liver tissue (after enzymatic hydrolysis).

Details on excretion:

Faecal excretion was the major route of elimination, and urinary excretion the secondary route.

Metabolite characterisation studies

Metabolites identified:

no

Details on metabolites:

Metabolites were not formally identified. The majority of bisphenol-AF was present in its conjugate form (> 86 %).

Bioaccessibility (or Bioavailability)

Bioaccessibility (or Bioavailability) testing results:

Not determined

Any other information on results incl. tables

Table 1       Concentration of bisphenol-AF in orally dosed rats

Tissues and excreta	Concentration (µg/kg)
	Without enzymatic hydrolysis				After enzymatic hydrolysis
	Rat 1	Rat 2	Rat 3	Rat 4	Rat 1	Rat 2	Rat 3	Rat 4
Liver	78.1	141.0	437.5	190.5	1210.2	1376.3	1637.5	1496.6
Muscle	8.1	10.0	12.8	17.1	44.3	20.1	17.6	19.5
Adipose	30.3	30.3	39.3	40.1	62.7	47.2	42.1	57.8
Testis	15.3	16.0	21.5	24.0	66.0	50.0	68.0	47.0
Kidney	31.4	28.3	56.3	45.5	880.3	387.0	372.5	431.3
Serum	3.4	3.2	3.9	4.3	1075.3	358.8	312.4	431.3
Urine	31.5	17.0	17.6	25.5	177.6	47.5	25.7	59.8
Faeces	424682.4	449908.6	337844.6	146468.7	545266.9	469864.7	370004.5	223646.9

Applicant's summary and conclusion

Conclusions:

Bisphenol-AF was poorly adorbed to the rat GI system where concentrations of test item greater than the application rate were observed in the faeces of individuals, indicating low potential for bioaccumulation. The highest level of bisphenol-AF were found in the liver, kidneys and serum. Extractions performed with and without enzyme hydrolysis indicated that the vast majority of recovered test item was present in its conjugated form, indicative of a test item that is readily metabolised by the liver. Faeces were confirmed as the major route of excretion of the test item with urine a secondary route.

Executive summary:

In a metabolism study bisphenol-AF (98 % purity), was administered to 4 male Sprague-Dawley rats over 14 consecutive days by oral gavage at a dose level of 10 mg/kg bw/day.

High levels of BPAF were detected in the liver, kidney and serum samples. The significant enhancement of the test item concentration after enzymatic hydrolysis in the serum, liver and kidney samples implies that the liver is the major organ responsible for metabolism and that the kidney plays an important part in the excretion of its metabolites. The highest level of bisphenol-AF was observed in the faeces, which indicates that most of the test item was excreted as the nonconjugated form. Faecal excretion was the major route of elimination, and urinary excretion was the secondary route.

Results were determined using a validated method where fortified samples, performed at three fortification levels, were extracted to confirm the sensitivity of the extraction and analytical methods. The mean recoveries ranged from 71.0 % to 102.3 % with relative standard deviations of no more than 13.2 % (n = 6) and the limit of quantification was determied to be 1 µg/kg for liver, muscle, adipose, kidney and urine samples and 3 µg/kg for faeces. Reported values indicate an adequately efficient and sensitive method.

This metabolism study in the rat is classified acceptable and satisfies the guideline requirement for a metabolism study OECD 417 in rats.