O-6-deoxy-α-L-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03/13/18 to 03/19/18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Vehicle:

yes

Details on test solutions:

Media preparation
The test medium used was OECD medium in accordance with OECD 201. It was reconstituted with chemical reagents and ultra-pure water.
Preparation date 23/03/2018
pH 7.05

Test solution preparation
Since the test item is poorly soluble under the test conditions, a saturated solution of test item (limit of solubility LS) was prepared. After a period of stirring (about 20 hours) to obtain maximum solubility, the aqueous phase was collected using 0,45 µm filters. Test solutions were be prepared by dilution of this solution.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

The Pseudokirchneriella subcapitata stock culture was maintained on sloped agar tubes at 4°C and regularly transferred to fresh medium at the laboratory. The algal stock culture was provided by the CCAP (Culture Collection of Algae and Protozoa), reference CCAP 1405/1.
A culture was prepared from the stock culture and maintained in an exponential growth by successive transfers every 3 to 4 days.

Test type:

static

Water media type:

other: ultra-pure water

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

Test item quantification was performed according to a validated analytical method a LC-MS/MS from Thermo (LC Ultimate 3000 and MS/MS TSQ Quantiva) as mass spectrometry detection to measure both test item concentrations at 0, 24, 48 and 72 hours.

Test temperature:

23°C

pH:

7.03

Details on test conditions:

Temperature: 22.7 to 23.3 °C
Light intensity: 8500 to 8600 lux.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Key result

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

ca. 68.18 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks:

(95 % confidence limits) (25.81 - 96.48 mg test item / L)

Key result

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

> 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 105.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Results with reference substance (positive control):

Potassium dichromate results:
CEt50 (mg.L-1) : 1.34
95% confidence limits (mg.L-1) : 1.25-1.42
Starting date : 26 October 2017

Validity criteria fulfilled:

yes

Remarks:

Validity criteria are met for both 48 and 72-hour endpoints

Conclusions:

The effects of O-6-deoxy-α-l-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate diluted in OECD test medium at the definitive rates of LS/16; LS/8; LS/4; LS/2 and LS (LS = limit of solubility) to Pseudokirchneriella subcapitata were investigated under laboratory conditions.
The study is valid since the growth multiplication factor in the control within the 48 and 72-hour test period was higher than 16 (actual values: 75 after 48h and 772 after 72h), the coefficient of variation of the average specific growth rates in the control was less than 7% (actual values:

2.74% after 48h and 1.82% after 72h) and the mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control was less than 35% (actual values: 3.07% after 48h and 5.03% after 72h). Since validity criteria were met after 48h, 48h results are presented below.
Based on growth rate, the NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the
EC50-48h were > 105.40 mg test item.L-1.

Based on yield rate, NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the EC50-
48h were > 105.40 mg test item.L-1.

Executive summary:

The objective of this study was to assess the effects of test item O-6-deoxy-α-l-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate on the growth of Pseudokirchneriella subcapitata over a period of 72 hours according to OECD 201 Guideline. Pseudokirchneriella subcapitata is a freshwater green algae that is recommended by the OECD Guideline 201.

Since the test item is poorly soluble under the test conditions, a saturated solution of test item (limit of solubility LS) was prepared. Test solutions were be prepared by dilution of this solution.

Test item quantification was performed according to a validated analytical method (final report RRCo-000146_01) LC-MS/MS from Thermo (LC Ultimate 3000 and MS/MS TSQ Quantiva) as mass spectrometry detection to measure O-6-deoxy-α-l-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate concentrations at 0, 24, 48 and 72 hours.

 

The effects of O-6-deoxy-α-l-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate diluted in OECD test medium at the definitive rates of LS/16; LS/8; LS/4; LS/2 and LS (LS = limit of solubility) to Pseudokirchneriella subcapitata were investigated under laboratory conditions.

The study is valid since the growth multiplication factor in the control within the 48 and 72-hour test period was higher than 16 (actual values: 75 after 48h and 772 after 72h), the coefficient of variation of the average specific growth rates in the control was less than 7% (actual values:

2.74% after 48h and 1.82% after 72h) and the mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control was less than 35% (actual values: 3.07% after 48h and 5.03% after 72h). Since validity criteria were met after 48h, 48h results are presented below.

Based on growth rate and on day by day geometric mean analysed concentrations, the NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the EC50-48h were > 105.40 mg test item.L-1.

 

Based on yield rate and on day by day geometric mean analysed concentrations, NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the EC50-48h were > 105.40 mg test item.L-1.

 

After 48h:

NOEC growth: 105.40 mg test item.L-1

NOEC yield: 105.40 mg test item.L-1

LOEC growth: > 105.40 mg test item.L-1

LOEC yield: > 105.40 mg test item.L-1

ECyield 10 (95% confidence limits): 68.18 mg test item.L-1 (25.81 – 96.48 mg test item.L-1)

ECyield 50 ECgrowth rate 10 ECgrowth rate 50: > 105.40 mg test item.L-1

Description of key information

The effects of O-6-deoxy-α-l-mannopyranosyl-(1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl (2α,3β,4α)-2,3,23-trihydroxyurs-12-en-28-oate diluted in OECD test medium at the definitive rates of LS/16; LS/8; LS/4; LS/2 and LS (LS = limit of solubility) to Pseudokirchneriella subcapitata were investigated under laboratory conditions.
The study is valid since the growth multiplication factor in the control within the 48 and 72-hour test period was higher than 16 (actual values: 75 after 48h and 772 after 72h), the coefficient of variation of the average specific growth rates in the control was less than 7% (actual values:

2.74% after 48h and 1.82% after 72h) and the mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control was less than 35% (actual values: 3.07% after 48h and 5.03% after 72h). Since validity criteria were met after 48h, 48h results are presented below.
Based on growth rate, the NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the EC50-48h were > 105.40 mg test item.L-1.

Based on yield rate, NOEC48h was 105.40 mg test item.L-1 and both the LOEC48h and the EC50-48h were > 105.40 mg test item.L-1.

Key value for chemical safety assessment

EC50 for freshwater algae:

105.4 mg/L