Oligomerisation and alkylation reaction products of 2-phenylpropene and phenol

Bioaccumulation: aquatic / sediment

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

bioaccumulation in aquatic species: fish

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study, basic data given (Jp. study report available with English results tables); test was performed by an laboratory with an high reputation for delivery of robust data.

Qualifier:

according to guideline

Guideline:

OECD Guideline 305 (Bioconcentration: Flow-through Fish Test)

Principles of method if other than guideline:

The test was performed by a laboratory with a high reputation for delivery of robust data.
This test was conducted in accordance with the test method "Bioaccumulation test of chemical substance in fish and shellfish" stipulated in the Order Prescribing the Items of the Test Relating to the New Chemical Substance (1974, Order of the Prime Minister, the Minister of Health and Welfare, the Minister of International Trade and Industry No.1). It corresponds to OECD TG 305 C of May 12, 1981)

GLP compliance:

no

Specific details on test material used for the study:

- Name of test material (as cited in study report or NITE database): 4-methyl-2,4-diphenylpent-1-ene or 2,4-diphenyl-4-methyl-1-pentene
- Dimer of methylstyrene and key constituent in OAPP
- Purity 97 % (according to data provided from client)
- No further information on test substance

Radiolabelling:

no

Details on sampling:

- Sampling intervals/frequency for test organisms: 2 fish every two weeks (0, 7, 14, 28, 42 and 60 days)
- Sampling intervals/frequency for test medium samples: twice a week
- Sample storage conditions before analysis: in Japanese
- Details on sampling and analysis of test organisms and test media samples (e.g. sample preparation, analytical methods): in Japanese

Vehicle:

yes

Details on preparation of test solutions, spiked fish food or sediment:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances):
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): 2-methoxyethanol and HCO-40
- Concentration of vehicle in test medium: high exposure level: 2-methoxyethanol ca. 25 ppm (v/v), HCO-40 0.2 mg/L; low exposure level: 2-methoxyethanol ca. 25 ppm (v/v), HCO-40 0.02 mg/L
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no data

Test organisms (species):

Cyprinus carpio

Details on test organisms:

TEST ORGANISM (see also CITI 1992)
- Common name: carp
- Source: Sugishima Fish Farm, Kumamoto, Japan
- Length at study initiation: 8 +/-4 cm
- Weight at study initiation: about 5 g
- Lipid content: 6.1 g (start); 7.3 g (end)
- Weight at termination: no data
- Method of breeding: flow through system at 24 ± 2°C
- Health status: only visibly healthy fish were used; after reception, fish were externaly desinfected (static conditions,
50 mg/L Terramycin (Taito Pfizer) and 7g/L sodium chloride for 24 h).
- Feeding during test:
- Food type: pelleted feed for carp (Japan Haigo Shiryo K.K.)
- Amount: 2% of total body weight
- Frequency: twice a day, half of total amount each

ACCLIMATION
- Acclimation period: at least one month
- Acclimation conditions (same as test or not): yes
- Type and amount of food: pelleted feed for carp (Japan Haigo Shiryo K.K.), 2% of total body weight
- Feeding frequency: twice a day, half of total amount each
- Health during acclimation (any mortality observed): no data, abnormal fish were removed.

Route of exposure:

aqueous

Test type:

flow-through

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

60 d

Total depuration duration:

16 d

Hardness:

--

Test temperature:

24 ± 2°C

Dissolved oxygen:

6 - 8 mg/L

Details on test conditions:

TEST SYSTEM (see also CITI 1992)
- Test vessel:
- Type (delete if not applicable): no data
- Material, size, headspace, fill volume: glas tank , volume 100 L
- Aeration: aeration of dilution water
- Type of flow-through (e.g. peristaltic or proportional diluter): no data
- Renewal rate of test solution (frequency/flow rate): 2.9 to 11.5 per day / flow rate 200 to 800 mL/min
- No. of organisms per vessel: 15 - 20
- No. of vessels per concentration (replicates): 1
- No. of vessels per control / vehicle control (replicates): no data
- Biomass loading rate: 4.5 - 6 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: underground water pumped at the site of Kurume Research Laboratories
- dilution water was confirmed to meet the quality criteria fisheries
- Intervals of water quality measurement: once every six month; temperature, pH and dissolved oxygen continuously.
- Intervals of test medium replacement: flow through system, see above

RANGE-FINDING / PRELIMINARY STUDY
- Results used to determine the conditions for the definitive study: test concentrations were decided on basis of an acute toxicity test (Oryzias latipes; 48h-LC50 5.4 mg/L)

Nominal and measured concentrations:

High exposure level (level 1): 10 µg/L
low exposure level (level 2): 1 µg/L

Reference substance (positive control):

no

Details on estimation of bioconcentration:

Calculation of bioconcentration factor = concentration of test substance in fish / concentration of test substance in water

Lipid content:

>= 3 - <= 7.7 %

Time point:

other: 7 d

Remarks on result:

other: Range of individual values: see Attached Document: BCF

Lipid content:

>= 1.2 - <= 4.5 %

Time point:

other: 60 d

Remarks on result:

other: Range of individual values: see Attached Document: BCF

Key result

Conc. / dose:

1 µg/L

Type:

BCF

Value:

2 767 L/kg

Basis:

whole body w.w.

Remarks:

lipid normalised (5 %) based on individual fish

Time of plateau:

14 d

Calculation basis:

steady state

Remarks on result:

other: average of measurements during steady state (three measurements from day 28 to day 60 with two fish per measurement)

Key result

Conc. / dose:

10 µg/L

Type:

BCF

Value:

2 320 L/kg

Basis:

whole body w.w.

Remarks:

lipid normalised (5 %) based on individual fish

Time of plateau:

14 d

Calculation basis:

steady state

Remarks on result:

other: average of measurements during steady state (three measurements from day 28 to day 60 with two fish per measurement)

Conc. / dose:

1 µg/L

Type:

BCF

Value:

2 912 L/kg

Basis:

whole body w.w.

Time of plateau:

28 d

Calculation basis:

steady state

Remarks on result:

other: average of measurements during steady state (three measurements from day 28 to day 60 with two fish per measurement)

Conc. / dose:

10 µg/L

Type:

BCF

Value:

1 790 L/kg

Basis:

whole body w.w.

Time of plateau:

14 d

Calculation basis:

steady state

Remarks on result:

other: average of measurements during steady state (three measurements from day 28 to day 60 with two fish per measurement)

Conc. / dose:

1 µg/L

Type:

BCF

Value:

>= 724 - <= 4 410 L/kg

Basis:

whole body w.w.

Time of plateau:

28 d

Calculation basis:

steady state

Remarks on result:

other: minimum and maximum value of three measurements with two fish per measurement during steady state from day 28 to day 60

Conc. / dose:

10 µg/L

Type:

BCF

Value:

>= 896 - <= 3 330 L/kg

Basis:

whole body w.w.

Time of plateau:

14 d

Calculation basis:

steady state

Remarks on result:

other: minimum and maximum value of three measurements with two fish per measurement during steady state from day 28 to day 60

Elimination:

yes

Parameter:

other: DT50 (high exposure)

Depuration time (DT):

4.5 d

Elimination:

yes

Parameter:

other: DT50 (low exposure)

Remarks:

probably mistaken (see re-estimation)

Depuration time (DT):

15.7 d

Remarks on result:

other: probably mistaken (see re-estimation)

Details on kinetic parameters:

The cumulated final tissue levels in whole fish on day 60 correlate well with either exposure level: 12.2 µg/g at 10 µg/L and 1.37 at 1 µg/L, i.e. a 10-fold higher exposure concentration resulted in a 10-fold higher tissue level. This indicates that the elimination kinetics at both exposure regimens are identical, which should end in similar eliminations constants or DT50 values. The presentation of the kinetic plots (Report, Fig. 13 and 14: see Attached Document BCF) suggest poor correlations between the residual fish levels over time, demonstrated by low regression coefficients R^2: R^2 (10 µg/L) = 0.58, R^2(1 µg/L) = 0.18 (rounded). In either case, the kinetics are obviously determined each by an outlier.
At least, the comparatively long DT50 of 15.7 d seems to be erroneous. A simple comparison of the initial tissue concentration (100%) with the residual one after 9 days suggest that the elimination half-lives are as follows (assuming first-order elimination kinetics):
High exposure (10 µg/L): decrease ~95%/9d = ~4 T/2 --> DT50 = ~2.3d
Low exposure (1 µg/L): decrease ~83%/9d = ~2.7 T/2 --> DT50 = ~3.3d.

Metabolites:

no data

Results with reference substance (positive control):

not included

Details on results:

Note: In the original study, BCF was normalised to a lipid content of 4% rather than 5% (not shown: see Attached Document BCF).

Reported statistics:

--

Original data (overview) (unadjusted to lipid content):

Day		7	14	28	42	60	BCF range from 28–60 days #
High	Conc. water(µg/l)	9.95	10.07	10.33	10.54	10.48
exposure group							896 -3330
	BCF replica1	1610	957	1370	896	1080
(10 µg/L)	BCF replica2	427	1950	2810	3330	1250
Low	Conc. water(µg/l)	1.01	0.98	0.94	0.94	0.93
exposure group							724 -4410
	BCF replica1	469	1610	3730	4410	724
(1µg/L)	BCF replica2	423	1740	2270	4120	2220

# Note: Original individual BCF values as measured, NOT normalised to a lipid content of 4 or 5%: Final reported BCF values have been related to 4% lipid (see Jp. Report, Table 10) . The values published in the NITE databank are misleading, because firstly neither has explicitly been stated that they cover the range of time-point specific BCFs, and because secondly nor is clear that they are original without lipid normalisation.

Single and mean BCFs after normalisation to 5% lipid content

Exposure group		42 d	60 d
10 µg/L	BCF replica 1	1659	1200
	BCF replica 2	2378	5208
	Average	2019	3204
1 µg/L	BCF replica 1	4690	905
	BCF replica 2	3075	2582
	Average	3883	1743

Validity criteria fulfilled:

not specified

Conclusions:

Despite lipid normalisation to 5 % , single BCF replicates were still relative variable in particular on day 60, with ratios between about 2.8 and 4.3. The highest mean BCFs are approx. 3200 (10 µg/L, 60 d) and 3900 (1 µg/L, 42 d). Averaged over a longer time period (still steady state), BCF are below 3000 L/kg. Based on these values, bioaccumulation is assessed to be moderate. Note: The authors concluded "low bioaccumulation" (NITE 2002: databank). Given the BCF < 2000 at 1 µg/L after 60 d, the bioconcentration potential is considered to be low, since -moreover- the lower experimental exposure concentration is closer to potential environmental water levels and therefore is of higher environmental relevance.

Reference 2

Endpoint:

bioaccumulation in aquatic species: fish

Type of information:

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

Adequacy of study:

weight of evidence

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The target substance oligomerisation and alkylation reaction products of 2-phenylpropene and phenol (OAPP) (EC no. 700-960-79) is obtained by a Lewis acid catalysed reaction between phenol and 1-methylstyrene. Products of the reaction are either methylstyrenated phenols or dimers/trimers of 1-methylstyrene (2-phenylpropene). Accordingly, the target substance OAPP consists of four basic groups of constituents. Two groups contain purely aryl-aliphatic (non-phenolic) substances differing only in the degree of oligomerisation (dimers and trimers of 2-phenylpropene). The third and fourth group contain mono-methylstyrenated and di-methylstyrenated phenols, respectively. Dimers and trimers amount together to about 45 to 80 % of OAPP, while the phenolic components contribute about 20 to 50 %.
The source substance p-cumylphenol (4-(1-methyl-1-phenylethyl)phenol) (CAS No. 599-64-4) is a main constituent of the target substance OAPP. It is a mono-methylstyrenated phenol present in OAPP and it is estimated that its bioaccumulation properties will also be representative for other phenolic components of OAPP.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance OAPP is a UVCB with a complex and variable composition of constituents. The source substance p-cumylphenol is a mono-constituent substance. It is a constituent of the target substance (mono-methylstyrenated phenol) thus contributing at least in part to the properties of the target substance.

3. ANALOGUE APPROACH JUSTIFICATION
Based on the chemical structure of the different components of OAPP, the bioaccumulation potential of the phenolic components is assessed to be lower than the bioaccumulation potential of non-phenolic components. Log Kow is somewhat lower compared to the same size non-phenolic components and the ability for biotransformation is considered to be better. Therefore, the source substance p-cumylphenol represents the part of the target substance OAPP that will bioaccumulate to a lesser extent. Bioconcentration factors determined for the source substance, represent the part of the target substance OAPP with a lower bioaccumulation potential.

Reason / purpose for cross-reference:

read-across source

Principles of method if other than guideline:

Read-across to the preceding entry:
Source substance: p-Cumylphenol (NITE, Japan, CSCL)_;
Reference: NITE National Institute of Technology and Evaluation, Japan 2002

Lipid content:

3.5 %

Time point:

start of exposure

Lipid content:

4.1 %

Time point:

end of exposure

Key result

Conc. / dose:

10 µg/L

Temp.:

25 °C

Type:

BCF

Value:

168 L/kg

Basis:

whole body w.w.

Time of plateau:

14 d

Calculation basis:

steady state

Remarks on result:

other: average of measurements during steady state (three measurements from day 28 to day 60 with two fish per measurement)

Remarks:

high exposure level

Conc. / dose:

10 µg/L

Temp.:

25 °C

Type:

BCF

Value:

>= 139 - <= 187 L/kg

Basis:

whole body w.w.

Time of plateau:

14 h

Calculation basis:

steady state

Remarks on result:

other: minimum and maximum value of three measurements with two fish per measurement during steady state from day 28 to day 60

Remarks:

high exposure level

Description of key information

The bioaccumulation potential of the substance Oligomerisation and alkylation reaction products of 2-phenylpropene and phenol (OAPP, previously phenol, methylstyrenated, Novares LA 300) has been assessed in bioaccumulation/ bioconcentration studies in fish. The BCF of two individual constituents of OAPP, one methylstyrenated (cumyl) substituted phenol and a dimer of 2-phenylpropene were experimentally shown to be low to moderate. For dimers and trimers of 2-phenylpropene, BMF were determined. Conversion into BCF indicates that the BCF for dimers is between 2000 and 5000, while the BCF for trimers is above 5000. However, the conversion of BMF into BCF values show such a high variability/standard deviation due to uncertainties in conversion methods that the high BCF values resulting from conversion have to be considered as not very reliable. They are not suited for assessment and should be taken with caution.

Key value for chemical safety assessment

BCF (aquatic species):

3 000 L/kg ww

BMF in fish (dimensionless):

0.137

Additional information

Aquatic bioaccumulation of OAPP and its constituents has been investigated in three bioconcentration/bioaccumulation studies. The constituents diphenylmethylpentene (4-methyl-2,4-diphenylpent-1-ene) (NITE 2002) and cumylphenol (1-methyl-1-phenylethyl)phenol (NITE 1990) were subject of valid bioconcentration flow-through fish tests (OECD 305). BCFs were determined to be 2767/2320 L/kg fish ww (low/high exposure concentration, lipid normalised) and 168 L/kg fish ww (high exposure concentration) at steady state, respectively. In a bioaccumulation study (dietary exposure, OECD 305-III) (Klix 2018), BMF factors could only be determined for the non-phenolic constituents of OAPP. For the phenolic components, the concentrations in fish during the depuration phase was too low to be determined. A bioaccumulation potential was thus found only for the non-phenolic constituents of OAPP. Lipid-based and growth-corrected BMF values were 0.0737 and 0.1374 for dimers and trimers, respectively. Conversion into BCF values by calculation of k₁rate constants using various methods resulted in average values of 2564 ± 1803 and 22338 ± 23353 (mean ± SD) for dimers and trimers, respectively. Their variation is very high and the results differ very much, so that especially the higher value is considered not to be reliable. This estimation of k₁ bears a high uncertainty indicated by the high standard deviation.

Overall, BCF are below 3000 for about 75 % (w/w) of the test substance (phenolic constituents and dimers) while for approx. 25 % (trimers) a reliable BCF value suited for assessment is not yet available. An estimate derived from the experimental BMF suggests that a BCF for trimers may be above 5000. Based on currently available evidence a preliminary BCF for OAPP is set at 3000 L/kg ww.