Trehalose

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:

The study will be available 31/08/2018

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)

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Name TREHALOSE, ANHYDROUS
Batch no. 160805
Appearance White crystalline powder
Composition Trehalose: 100 % (dry basis)
Production date 05. Aug. 2016
Expiry date 05. Aug. 2018
Storage Room temperature (20 ± 5 °C), Keep away from humidity

Analytical monitoring:

yes

Details on sampling:

Test item solutions with concentration of 100 mg/L (nominal) were diluted twofold with medium to achieve concentrations in the calibrated range (5 – 50 mg/L).

Vehicle:

no

Details on test solutions:

A stock solution containing 100.0 mg/L in algal medium (demineralised water enriched with minerals but without algae) was prepared. The lower treatments will be prepared by dilution of this stock solution with algal medium

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Unicellular freshwater green alga.
Genus Desmodesmus
Species subspicatus
SAG Strain Number 86.81
Taxonomic position Chlorophyta - Chlorophyceae

The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen). The algae are kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture was prepared. From an aliquot of the permanent culture, the pre-culture was prepared.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

no

Hardness:

The algal medium corresponds to the OECD TG 201 medium.

Test temperature:

22.5 – 24.3 °C

pH:

The algal medium corresponds to the OECD TG 201 medium.
measured 7.8 - 9

Dissolved oxygen:

The algal medium corresponds to the OECD TG 201 medium.

Salinity:

The algal medium corresponds to the OECD TG 201 medium.

Conductivity:

The algal medium corresponds to the OECD TG 201 medium.

Nominal and measured concentrations:

4.6 / 10 / 22 / 46 / 100 mg/L nominal concentration
5.4 / 7.9 / 10.2 / 14.3 / 20.4 mg/L Geometric Mean

In the first experiment the measured concentrations lay between 101 % and 123 % of the nominal concentrations at the beginning of the test. In the second experiment the meas-ured concentrations lay between 83 % and 128 % of the nominal concentrations at the beginning of the test. In both experiments the test solution became turbid until the end of the test and after 72 h incubation no test sample could be detected. Therefore, the geo-metric mean is used for statistical evaluation. The geometric mean is formed from the ini-tial values and the LOD (5 mg/L).
Geometric mean is calculated by multiplication of the n participating concentrations and taking the nth root.
The concentration of the test item in the test vessels was measured at the start and at the end of the test. An additional replicate without algae for each test concentration and the control was prepared only for analytical measurement.

Details on test conditions:

Date: 25. – 29. Jun. 2018
Treatments tested: 4.6 / 10 / 22 / 46 / 100 mg/L nominal concentration
The concentrations to be tested are based on non GLP pre-tests
Number of replicates: 6 replicates for the blank control
3 replicates for each treatment
1 replicate for each test concentration and the control without alga only for analytical measurement
Vessels: glass flasks total volume 65 mL
Duration: 72 hours
Lighting: 5000 lux
Temperature: 22.5 – 24.3 °C
Replicates (Blank control): 45 ±1 ml algal medium and alga
Replicates (Treatments): 45 ±1 ml test solution and alga

Reference substance (positive control):

yes

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

5.42 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

5.42 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield = Cell concentration /mL

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

7.86 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

7.86 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield = Cell concentration /mL

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

12.51 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

6.43 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield = Cell concentration /mL

Key result

Duration:

72 h

Dose descriptor:

EC50

Remarks:

value is extrapolated

Effect conc.:

30.41 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

13.54 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Yield = Cell concentration /mL

Details on results:

In both experiments the test solution became turbid until the end of the test and after 72 h incubation no test sample could be detected

Results with reference substance (positive control):

The 72h-EC50 values of potassium dichromate (K2Cr2O7, CAS No. 7778-50-9) were deter-mined in a separate reference test. The values lay within the range of the laboratory (growth rate 0.73 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

Reported statistics and error estimates:

Calculation of results was performed with the help of validated software (Microsoft Ex-cel®). The estimation of the biological data was accomplished using the software ToxRat® Professional, version 3.2.1.

Validity criteria fulfilled:

yes

Conclusions:

Under the test conditions, Trehalose showed effects on Algae and following toxicological dose discriptors are derived:
Endpoint NOEC LOEC EC10 EC50
Growth Rate 5.42 mg/L 7.86 mg/L 12.51 mg/L 30.41 mg/L (extrapolated)
Yield 5.42 mg/L 7.86 mg/L 6.43 mg/L 13.54 mg/L

Executive summary:

Two experiments were performed. In the 1^stexperiment one of the validity criteria was not met. Therefore, the experiment was repeated.Nevertheless, the inhibition values and the measured test item concentration in both experiments was in the same range. With the exception of the measured test item concentrationthe results of the 1^stexperiment are not stated in the report, but will be kept together with the other raw data in the GLP archive of the test facility.

 

The study was performed using 5 concentrations ranging from 4.6 to 100 mg/L. Incubation time (test systemDesmodesmus subspicatus)was 72 hours. The cell concentration of each replicate was determined by measuring the cell numbers every 24 hours with an electronic particle counter. Growth rate µ and the yield[1]were determined from the cell number at the respective observation times.

 

Significant inhibition of algal growth was observed at the following concentrations:

10 / 22 / 46 / 100 mg/L.

 

In treatment 22 mg/L the algae cells were coagulated and measurement of the cell number therefore biased. The calculated inhibition values in this treatment were implausible low. Therefore, this treatment was not used for evaluation.

At the start and at the end of the test, the content of the test item in the test solutions (test solutions without algae) was determined using HPLC.

Deviating from analytical measurement in a daphnia and a fish test, no test item can be detected at the end of the test. This effect, insufficient stability of the test item under algal test conditions was observed in both experiments.

Therefore, the geometric mean is used for statistical evaluation. The geometric mean is formed from the initial values and the LOD (5 mg/L).

 

The 72h-EC₅₀values of potassium dichromate (K₂Cr₂O₇, CAS No. 7778-50-9) were determined in a separate reference test. The values lay within the range of the laboratory (growth rate 0.73 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

The following results for the test itemTREHALOSE, ANHYDROUSwere determined:

Results of the test item

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	5.42 mg/L	7.86 mg/L	12.51 mg/L	30.41 mg/L (extrapolated)
Yield	5.42 mg/L	7.86 mg/L	6.43 mg/L	13.54 mg/L

[1]Yield (according to OECD Guideline 201) is defined as the biomass at the end of the exposure period minus the biomass at the start of the exposure period. Calculation see under 8.2 Growth and growth inhibition are quantified from measurements of the algal biomass as a function of time. Algal biomass is defined as the dry weight per volume, e.g. mg algae/L test solution. However, dry weight is difficult to measure and therefore surrogate parameters are used. Of these surrogates, cell counts are most often used.

Description of key information

one toxicity study available.

Due to reaction of the substance with the used media, the geometric mean of measured concentrations is used for determination of EC50.

As all other ecotoxicological tests (aquatic invertebrates, Fish embryo toxicity) indicate no hazard, the substance is readily biodegradable and has a Log Pow < 0.3, it is judged that the findings of the algae study do not indicate an ecotoxicological hazard.

Key value for chemical safety assessment

EC50 for freshwater algae:

30.41 mg/L

EC10 or NOEC for freshwater algae:

12.51 mg/L