3,5-dihydroxy-2,6,6-tris(3-methylbuten-2-yl)-4-(3-methyl-1-oxobutyl)cyclohexa-2,4-dien-1-one

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

48 hours

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

The study performed on hop extract is relevant for beta acids, since hop extract may contain up to ca. 64% beta acids, and the extract tested here contained 38% beta acids. A further study has been planned for beta acids and will be entered as an update.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Batch 160025

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

Media was stirred for 22 hours and 15 minutes and settled for 4 hours. After settling the first 100ml (approximately) of aqueous phase was removed (avoiding all settled and floating material) and discarded. The remaining aqueous phase provided sufficient volumes for water quality
measurements and testing.

Test organisms (species):

Daphnia magna

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

pH:

7.08 - 7.57

Dissolved oxygen:

6.17 - 7.31 mg/l

Nominal and measured concentrations:

Loading rates 0.1, 1.0, 10 and 100mg/l from stock solutions (WAF) used in preliminary study
Loading rates 1.0, 1.8, 3.2, 5.8 and 10.4mg/l were used in the main study

Reference substance (positive control):

yes

Remarks:

1.4 mg/l potassium dichromate in a separate GLP study

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 2.242 mg/L

Nominal / measured:

nominal

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 5.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat. (total fraction)

Basis for effect:

mobility

In range finder, 1 mg/l loading lead to no effect, but at 10 mg/l and 100 mg/l, there was 100% immobilisation

In main study, 2.42 or lower concentrations gave no adverse effects

A concentration of 5.8 mg/l resulted in 30% (6/20) immobilisation

A concentration of 10 mg/l resulted in 0% immobilisation (possibly due to partition effects from undissolved material)

Validity criteria fulfilled:

yes

Conclusions:

The 24-hour EC50 and 48-hour EC50 of HOP EXTRACT to Daphnia magna were >2.24mg/l and >2.24mg/l respectively (both determined by Linear Interpolation).
The 0 to 24-hour NOEC and LOEC were 2.242mg/l and >2.24mg/l respectively
The 0 to 48-hour NOEC and LOEC were 2.242mg/l and >2.24mg/l respectively
The Daphnia were examined at 24-hours and 48-hours for abnormal behaviour during the determination of immobility. All Daphnia appeared normal with no signs of abnormality
No test concentration immobilised 100% Daphnia and the highest concentration where no immobilisation occurred was 2.242mg/l.

Executive summary:

The study performed on hop extract is relevant for beta acids, since hop extract may contain up to ca. 64% beta acids, and the extract tested here contained 38% beta acids. A further study has been planned for beta acids and will be entered as an update.

Higher treatment levels up to 10 mg/l were also tested, but at higher concentrations, there was a lower rate of immobilisation, perhaps due to partitioning of biologically active components into undissolved oils.

The water solubility has been hard to determine due to the multiple components in the substance; the biological effects seen in this study indicate that there is dissolved material present

Reference 2

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

48 hours

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Batch DB EXP 10

Analytical monitoring:

yes

Vehicle:

yes

Remarks:

Dilution water

Details on test solutions:

Information provided by the Sponsor indicated that Potassium salt of Beta acids partially
dissolves in aqueous alkaline solutions. Beta acids dissolve up to 7.7g/litre in water with the other
constituents remaining undissolved. The sample was therefore tested as insoluble.
Both the preliminary range-finding test and the definitive test were prepared by the direct addition
of measured amounts of test substance to dilution water and stirred.

The only practicable method to prepare the 3, 6 and 12mg/l test concentration was by dilution
(addition of 31.25, 62.5 and 125ml of the 24mg/l preparation to 250ml of dilution water
respectively). The solution then was stirred for 24 hours and 45 minutes and settled for 4 hours.
After settling the first 100ml (approximately) of aqueous phase was removed (avoiding all settled
and floating material) and discarded. The remaining aqueous phase provided sufficient volumes for
water quality measurements and testing.

Test organisms (species):

Daphnia magna

Test type:

static

Water media type:

freshwater

Total exposure duration:

48 h

Test temperature:

19.8 - 20.9 oC

pH:

6.89 - 7.58

Dissolved oxygen:

6.89 - 7.39 mg/l

Nominal and measured concentrations:

A preliminary (range-finding) test was conducted at
concentrations of 0 (control), 0.1, 1, 10 and 100mg/l.
The duration of the preliminary test was 48 ± 1 hours.
There was a single replicate at each concentration.
Preliminary test results: Data from the preliminary test identified the 48-hour EC50
as being between 10-100mg/l.

Definitive test:
Number of replicates: Four replicates at each concentration.
Nominal concentrations:
0 (control), 3, 6, 12, 24, and 48 mg/l
Mean measured concentrations:
0 (control), 1.542, 1.779, 2.796, 4.486, 6.708 mg/l

Reference substance (positive control):

yes

Remarks:

1.4 mg/l potassium dichromate in a separate GLP study

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 1.542 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr. (dissolved fraction)

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 1.87 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr. (dissolved fraction)

Basis for effect:

mobility

Data from the preliminary test identified the 48-hour EC50

as being between 10-100mg/l.

The definitive test conducted from 31 July to 02 August 2018 was performed according to the

OECD 202 2004 guideline and met all validity criteria.

The 24-hour EC50 and 48-hour EC50 of Potassium salt of Beta acids to Daphnia magna were 1.87

mg/l and 1.87 mg/l respectively (determined by Maximum Likelihood-Probit). Graphical

representations of the 24-hour and 48-hour EC50 values are given in Graphs 1 and 2 respectively.

The 0 to 24-hour NOEC and LOEC were 1.542 mg/l and 1.779 mg/l respectively (determined by

Steel's Many-One Rank Test). The 0 to 48-hour NOEC and LOEC were 1.542 mg/l and 1.779

mg/l respectively (determined by Steel's Many-One Rank Test). A graphical representation of the

48-hour dose response plot is given in Graph 3.

The Daphnia were examined at 24-hours and 48-hours for abnormal behaviour during the

determination of immobility. All Daphnia appeared normal with no signs of abnormality.

The lowest test concentration which immobilised all twenty Daphnia after 48-hours was 2.796

mg/l. Immobilisation was seen in all test concentrations.

None (0%) of the twenty control Daphnia maintained in dilution water were immobilised and none

were trapped in the meniscus during the study. This fulfills the control validity criteria of the study

as the percentage of immobilisation did not exceed 10%, confirming that the test organisms were of

suitable “health” for use in the study.

Validity criteria fulfilled:

yes

Conclusions:

The definitive test conducted from 31 July to 02 August 2018 was performed according to the
OECD 202 2004 guideline and met all validity criteria.
The 24-hour EC50 and 48-hour EC50 of Potassium salt of Beta acids to Daphnia magna were 1.87
mg/l and 1.87 mg/l respectively (determined by Maximum Likelihood-Probit). Graphical
representations of the 24-hour and 48-hour EC50 values are given in Graphs 1 and 2 respectively.
The 0 to 24-hour NOEC and LOEC were 1.542 mg/l and 1.779 mg/l respectively (determined by
Steel's Many-One Rank Test). The 0 to 48-hour NOEC and LOEC were 1.542 mg/l and 1.779
mg/l respectively (determined by Steel's Many-One Rank Test). A graphical representation of the
48-hour dose response plot is given in Graph 3.
The Daphnia were examined at 24-hours and 48-hours for abnormal behaviour during the
determination of immobility. All Daphnia appeared normal with no signs of abnormality.
The lowest test concentration which immobilised all twenty Daphnia after 48-hours was 2.796
mg/l. Immobilisation was seen in all test concentrations.
None (0%) of the twenty control Daphnia maintained in dilution water were immobilised and none
were trapped in the meniscus during the study. This fulfills the control validity criteria of the study
as the percentage of immobilisation did not exceed 10%, confirming that the test organisms were of
suitable “health” for use in the study.

Executive summary:

EC50 (48h): 1.87 mg/L

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

1.87 mg/L