Reaction mass of stearoyl sarcosine and palmitoyl sarcosine

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

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

Remarks:

Summary of available data used for the endpoint assessment of the target substance

Adequacy of study:

key study

Justification for type of information:

see category justification attached to chapter 13

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

1 300 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other:

Remarks:

Source: EC 701-177-3, Schill+Seilacher, 2009, LAUS, 09070901G701, activated sludge

Duration:

3 h

Dose descriptor:

other: EC20

Effect conc.:

130 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other:

Remarks:

Source: EC 701-177-3, Schill+Seilacher, 2009, LAUS, 09070901G701, activated sludge

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other:

Remarks:

Source: EC 701-177-3, Schill+Seilacher, 2009, LAUS, 09070901G701, activated sludge

Duration:

3 h

Dose descriptor:

IC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other:

Remarks:

Source: EC 701-177-3, BASF, 1989, Ciba Geigy, 894071, activated sludge

Duration:

3 h

Dose descriptor:

other: IC20

Effect conc.:

75 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other:

Remarks:

Source: EC 701-177-3, BASF, 1989, Ciba Geigy, 894071, activated sludge

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 Sep 2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

adopted in 1984

Deviations:

yes

Remarks:

Higher temperature during the test (Guideline: 18 - 20 °C, Test: 23.3 - 24.4 °C)

Qualifier:

according to guideline

Guideline:

EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)

Version / remarks:

adopted in 2008

GLP compliance:

yes (incl. QA statement)

Remarks:

Landesamt für Umwelt, Wasserwirtschaft und Gewerbeaufsicht, Mainz, Germany

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution containing 10 g/L was prepared using the following procedure: 10 g test item was mixed with 1.1 g NaOH (45%). 40 mL deionised water (80 – 100 °C) was added. After homogenisation with a glass stirrer, 85 mL deionised water (room temperature) was added. Then, the vessel was stirred on a magnetic stirrer. 0.5 M NaOH was added until the resulting solution was clear. The pH of the solution was adjusted to 7.8 ± 0.2 with 1 M HCl. the solution was transferred into a volumetric flask and filled up to 1000 mL with deionised water.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Source of the inoculum: The sludge was taken from the activation basin of the sewage plant in Edenkoben, Germany
- Laboratory culture: Upon arrival, the sludge was washed with tap water three times, centrifuged and resuspended in tap water.
- Suspended solids: 4.2 g/L

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

pH:

7.7 - 8.1

Nominal and measured concentrations:

Nominal: 0, 1.0, 3.2, 10, 32, 100, 320, 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Beakers
- Material, fill volume: Glass, 500 mL
- Aeration: Purified air at 0.75 L/min
- No. of vessels per concentration (replicates): 2 for treatment 1000 mg/L, 1 for all other treatments and positive control
- Concentration of suspended solids: 1.7 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water
- Total organic carbon: < 1.0 mg/L
- Metals (mg/L): Na 4.7, K 2.6, Ca 33.1, Mg 2.1, other metals < 0.0005 to < 0.01
- Pesticides: < LOD
- Chloride: 9.0 mg/L
- Conductivity: 189 µS/cm
- Intervals of water quality measurement: pH was measured at test end.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable)
- Respiration rate was measured after 3 h of incubation

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Duration:

3 h

Dose descriptor:

other: EC20

Effect conc.:

130 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: 95% CL = 62 - 310 mg/L

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

1 300 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: 95% CL = 480 - 7200 mg/L; extrapolated

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- The inhibition caused by the test item did not rise above 45%. Therefore the EC50(3h) was determined by extrapolation.
- All validity criteria are fulfilled.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50: 9.1 mg/L, 95% CL = 7.2 - 11.0 mg/L

Table 1: Inhibition (%) of microbial respiration after 3 h of incubation

Concentration	% Inhibition
Control	-
1.0	-3.2
3.2	1.2
10	1.7
32	16.9
100	25.5
320	24.0
1000 (1)	43.8
1000 (2)	45.3

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

08 Mar 1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

adopted in 1984

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test item was directly added to the test medium.

Test organisms (species):

activated sludge

Details on inoculum:

- Source of inoculum: Activated sludge was collected from the sewage treatment plant of Therwil, Switzerland on 06 Mar 1989.
- Preparation of inoculum for exposure: The sludge was separated from the aqueous layer by settling.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Test temperature:

18 - 20 °C

Nominal and measured concentrations:

Nominal: 0, 1.1, 3.3, 10.7, 32.0, 100.2 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: BOD flasks with gas inlet
- Size, fill volume: 250 mL, 200 mL
- Aeration: Yes
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2
- Concentration of sludge: 1.6 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dechlorinated drinking water

EFFECT PARAMETERS MEASURED (with observation intervals if applicable)
- Microbial respiration was measured after 3 h of incubation.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: approx. 3.2

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Duration:

3 h

Dose descriptor:

other: IC20

Effect conc.:

75 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

IC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Validity criteria are fulfilled.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: IC50 = 13 mg/L

Table 1: Inhibition of microbial respiration

Concentration (mg/L)	Consumption Rate (mg/L*h)	Inhibition (%)
Blank 1	40.62	2
Blank 2	42.21	-2
1.1	41.62	0
3.3	43.97	-6
10.7	41.79	-1
32.0	35.70	14
100.2	30.00	28

Description of key information

NOEC (3 h) = 10 mg/L (OECD 209, activated sludge); read-across

Additional information

No studies investigating the toxicity to microorganisms are available for Reaction mass of N-(1-oxooctadecyl)sarcosine and N-hexadecanoyl-N-methylglycine (EC 947-850-7). Therefore, in accordance with Regulation (EC) No. 1907/2006 Annex XI, 1.5 a read-across to the structurally related source substance N-methyl-N-(C18-(unsaturated)alkanoyl)glycine (EC 701-177-3) was conducted.

The chemical properties of the target and source substance are similar due to the structural similarities, i.e., sarcosine head group and the hydrocarbon tail. In addition, due to the pka of the target substance (pKa = 3.96, major constituent), at physiological pH values (above 6) the target substance is expected to be in the anionic form, as are the source substances. As a result, the chemistry of the target and source substance, in solution (above pH 6), are expected to be similar. Thus, the source substance is considered suitable representative for the evaluation of the toxicity to microorganisms of the target substance. The read-across approach is justified in detail within the analogue justification in IUCLID section 13.

The key study conducted with the source substance N-methyl-N-(C18-(unsaturated)alkanoyl)glycine (EC 701-177-3) was carried out under GLP conditions according to OECD guideline 209. Effects on the respiration rate of microorganisms were observed at concentrations of 32 mg/L and above (nominal), leading to an estimated EC50 (3 h) of 1300 mg/L (extrapolated) and a NOEC (3 h) of 10 mg/L.

The supporting study conducted with the source substance N-methyl-N-(C18-(unsaturated)alkanoyl)glycine (EC 701-177-3) according to OECCD guideline 209 did not follow GLP standards. Five test concentrations (1.1, 3.3, 10.7, 32.0, 100.2 mg/L) were inoculated with activated sludge. In this study, the IC50 (3 h) for respiration inhibition was above the highest tested concentration of 100 mg/L.

Based on the available results from a structurally related read-across substance (in accordance to Regulation (EC) No 1907/2006 Annex XI, 1.5) which is characterized by a comparable structure and chemistry, it can be concluded that Reaction mass of N-(1-oxooctadecyl)sarcosine and N-hexadecanoyl-N-methylglycine (EC 947-850-7) will exhibit comparable effects on activated sludge microorganisms.