Saccharomyces cerevisiae, ext.

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to OECD guidelines and in compliance with GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L)

IUPAC Name:

Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L)

Constituent 2

Reference substance name:

Saccharomyces cerevisiae, ext.

EC Number:

283-294-5

EC Name:

Saccharomyces cerevisiae, ext.

Cas Number:

84604-16-0

Molecular formula:

Not applicable as the substance is an UVCB

IUPAC Name:

Yeast extract, Saccharomyces cerevisiae

Details on test material:

Description Light beige powder
Batch 071002230
Purity Not indicated by the sponsor; treated as 100% pure
Test substance storage At room temperature in the dark
Stability under storage conditions Stable
Expiry date 31 May 2013

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Stability in water Not indicated
Solubility in water Completely

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: control and the limit concentration(100 mg/l)
- Sample storage conditions before analysis: keept in refrigerator

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Preparation started with a concentration of 100 mg/l. No other treatment than vigorous shaking was needed to completely dissolve the test substance in the test medium. The lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. The highest concentration was clear and very slightly yellow, whereas all other test solutions were all clear and colourless.

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name:Pseudokirchneriella subcapitata,
- Strain: strain: NIVA CHL 1
- Source (laboratory, culture collection): In-house laboratory culture.
- Age of inoculum (at test initiation): 3 days

ACCLIMATION
- Culturing media and conditions (same as test or not): No, culture M1, test M2
- Any deformed or abnormal cells observed: no

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Remarks on exposure duration:

no

Post exposure observation period:

none

Test conditions

Hardness:

0.24 mmol/l (24 mg CaCO3/l)

Test temperature:

The temperature of the test medium was 23.0°C at the start of the test. During the exposure period the temperature measured in the incubator was maintained between 22.8 and 23.7°C. Temperature remained within the limits prescribed by the protocol (21-24°C, constant within 2°C).

pH:

7.6 - 8.0

Dissolved oxygen:

not measured

Salinity:

no data

Nominal and measured concentrations:

Limit test:
nominal: 100 mg/l , measured (Time Weight Average concentration) 19 mg C/l (measured as TOC

Range finding test:
nominal: 0.1, 1.0 and 10 mg/l

Details on test conditions:

Test duration 72 hours
Test type Static
Test vessels 100 ml, all-glass, containing 50 ml of test solution
Medium M2
Cell density An initial cell density of 1 x 104 cells/ml.
Illumination Continuously using TLD-lamps of the type ‘Cool-white’ of 30 Watt, with a light intensity within the range of 84 to 91 uE.m-2.s-1.
Incubation Vessels were distributed at random in the incubator. During incubation the algal cells were kept in suspension by continuous shaking.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 19 other: mg C/L

Nominal / measured:

meas. (TWA)

Conc. based on:

other: Total Organic Carbon

Basis for effect:

growth rate

Effect concentrations 2

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

19 other: mg C/L

Nominal / measured:

meas. (TWA)

Conc. based on:

other: Total Organic Carbon

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes (factor 93)

No significant differences were recorded between the values for growth rate or yield at the limit concentration when compared to the control group. Note that at the concentration of 10 mg/l growth was and yield were increased rather than reduced/inhibited.

Microscopic observations at the end of the test revealed a normal and healthy appearance of the exposed cells when compared to the control.

Results with reference substance (positive control):

- Results with reference substance valid? yes
Potassium dichromate reduced growth rate of this fresh water algae species at nominal concentrations of 0.56 mg/l and higher.

The EC50 for growth rate reduction (ERC50: 0-72h) was 1.1 mg/l with a 95% confidence interval ranging from 1.0 to 1.3 mg/l. The historical ranges for growth rate reduction lie between 0.82 and 2.3 mg/l. Hence, the ERC50: 0-72h for the present batch corresponds with this range.

The EC50 for yield inhibition (EYC50: 0-72h) was 0.59 mg/l with a 95% confidence interval ranging from 0.51 to 0.68 mg/l. The historical ranges of the 72h EC50 for yield inhibition lie between 0.43 and 1.1 mg/l. Hence, the EYC50: 0-72h for the present batch corresponds with this range.

Reported statistics and error estimates:

no data

Any other information on results incl. tables

Mean cell densities (x 10⁴cells/ml)

Nominal conc.	Exposure time (hours)
test substance^1
(mg/l)	0	24	48	72
control	1.0	4.4	36.9	92.8
0.10	1.0	4.0	36.1	86.5
1.0	1.0	3.9	30.5	88.5
10	1.0	4.2	25.3	145.3
100 (19)	1.0	2.0	13.9	95.6

1.Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L)

() – between brackets the Time Weight Average concentration is given.

Percentage reduction of growth rate (total test period) and percentage inhibition of yield

Nominal conc. test substance^1 (mg/l)	Mean growth rate		Yield (0-72 h)
	µ (0-72 h)	Reduction^2 (%)	x10^4cells/ml	Inhibition^2(%)
control	0.06290		91.75
0.10	0.06195	1.5	85.50	6.8
1.0	0.06223	1.1	87.50	4.6
10	0.06914	-9.9	144.33	-57.3
100 (19)	0.06324	-0.5	94.58	-3.1

  1.Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L)

2. Negative numbers indicate an increase of growth/yield.

() – between brackets the Time Weight Average concentration is given.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

No reduction of growth rate or inhibition of yield was recorded at the limit concentration of Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L) tested.

The EC50 for both growth rate reduction (ERC50: 0-72h) and the EC50 for yield inhibition (EYC50: 0-72h) was beyond the range tested, i.e. exceeded a TWA concentration of 19 mg C/l.

The NOEC for growth rate reduction and yield inhibition equalled the TWA concentration of 19 mg C/l.

Executive summary:

The batch of Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L) tested was a light beige powder with unknown purity. The test substance was completely soluble in test medium at the concentrations tested.

 Preparation of test solutions started with a concentration of 100 mg/l. No other treatment than vigorous shaking was needed to completely dissolve the test substance in the test medium. The lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium.

 A combined limit/range-finding test was performed. Six exponentially growing algal cultures per group were exposed to a control and a concentration of 100 mg/l. Additionally, three replicates per concentration were exposed to 0.1, 1.0 and 10 mg Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L) per litre. The initial cell density was 10⁴cells/ml. The total exposure time was 72 hours and the samples for the analytical confirmation of actual exposure concentrations were taken at the start, after 24 hours of exposure and at the end of the test period.

Analyses were based on Total Organic Carbon (TOC) contents. The actual measured concentration in the limit concentration at the start of the test was 36 mg C/l when corrected for the carbon measured in the control samples. The measured concentration decreased to 22 mg C/l after 24 hours of exposure and further to 10 mg C/l after 72 hours of exposure. The corresponding concentrations of Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L) were 94, 56 and 25 mg/l, respectively. Based on these results the Time Weight Average concentration was 19 mg C/l, which corresponds to 49 mg/l of Saccharomyces cerevisiae, Extract (Springer 0207/0-MG-L).

The study met the acceptability criteria prescribed by the protocol and was considered valid.

The EC₅₀for both growth rate reduction (E_RC₅₀: 0-72h) and the EC₅₀for yield inhibition

(E_YC₅₀: 0-72h) was beyond the range tested, i.e. exceeded a TWA concentration of 19 mg C/l, which corresponds to 49 mg/l ofSaccharomyces cerevisiae, Extract (Springer 0207/0-MG-L).

The NOEC for growth rate reduction and yield inhibition equalled a TWA concentration of

19 mg C/l, which corresponds to 49 mg/l ofSaccharomyces cerevisiae, Extract (Springer 0207/0-MG-L).