Manganese sulphide

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2010-02-11 to 2010-02-12

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP and a current guideline.

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Guideline:

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

Guideline:

EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

For the purpose of the test, the test item was dispersed in water. Amounts of test item (5, 16, 50, 160 and 500 mg) were each separately dispersed in approximately 250 mL of water and subjected to ultrasonication for approximately 15 minutes followed by magnetic stirring for 24 hours in order to maximise the dissolved test item concentration. All test vessels were shielded from the light during mixing. Synthetic sewage (16 mL), activated sewage sludge (200 mL) and water were added to a final volume of 500 mL to give the required concentrations of 10, 32, 100, 320 and 1000 mg/L.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Source of inoculum: A mixed population of activated sewage sludge micro-organisms was obtained on 12 February 2010 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK which treats predominantly domestic sewage.

- Method of cultivation: The activated sewage sludge sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21ºC and was used on the day of collection.

- Preparation of inoculum for exposure: Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 mL) of the activated sewage sludge by suction through a pre-weighed GF/A filter paper using a Buchner funnel which was then rinsed 3 times with 10 mL of deionised reverse osmosis water and filtration continued for 3 minutes. The filter paper was then dried in an oven at approximately 105ºC for at least 1 hour and allowed to cool before weighing. This process was repeated until a constant weight was attained.

- Initial biomass concentration: The suspended solids concentration was equal to 4.0 g/L prior to use.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

140 mg/L as CaCO3

Test temperature:

21±1°C

pH:

see Table 2.

Salinity:

not reported

Nominal and measured concentrations:

Test based on nominal concentrations of 10, 32, 100, 320 and 1000 mg/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: 500 mL conical flask
- Aeration: aerated with clean, oil-free compressed air via narrow bore glass tubes at a rate of approximately 0.5 – 1 litre per minute

TEST MEDIUM / WATER PARAMETERS
- Source of dilution water: ZDB13 ALVASTON
- Preparation of dilution water: The test water was dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex water softener). After dechlorination and softening the water was then passed through a series of computer controlled plate heat exchangers to achieve the required temperature.
- Total organic carbon: 1.018 mg/L
- Pesticides: 0.001 µg/L
- Chlorine: 0.307 mg/L
- Alkalinity: 7.6
- Ca/mg ratio: not reported
- Conductivity: 386.192 µS/cm at 20°C

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod/ Light intensity: normal laboratory lighting

EFFECT PARAMETERS MEASURED: The rate of respiration was determined after 3 hours contact time with the test item

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.1 - 3.2
- Test concentrations: 10, 32, 100, 320 and 1000 mg/L (nominal)

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 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:

NOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

Oxygen consumption rates and percentage inhibition values for the control, test and reference items are given in Table 1. The pH values of the test preparations at the start and end of the exposure period are given in Table 2, and observations made on the test preparations throughout the study are given in Table 3.

Observations made at 0 hours, 30 minutes and 3 hours (Table 3) showed that the control vessels contained a dark brown dispersion and the reference item vessels contained a dark brown dispersion with no undissolved reference item visible.
Observations of the test item vessels made at 0 hours prior to the addition of activated sewage sludge and synthetic sewage showed that the test concentration of 10 mg/L contained an extremely pale brown dispersion with brown particles of test item dispersed throughout and the test concentration of 32 mg/L contained a pale brown dispersion with brown particles of test item dispersed throughout. The test concentration of 100 mg/L contained a brown dispersion with brown particles of test item dispersed throughout and the test concentration of 320 mg/L contained a dark brown dispersion with brown particles of test item dispersed throughout. The test concentration of 1000 mg/L contained a very dark brown dispersion with brown particles of test item dispersed throughout.
Observations of the test item vessels made after 30 minutes and 3 hours contact time showed that the test concentrations of 10 and 32 mg/L contained a dark brown dispersion with few particles of test item visible dispersed throughout. The test concentrations of 100 and 320 mg/L contained a dark brown dispersion with particles of test item visible dispersed throughout. The test concentration of 1000 mg/L contained a very dark brown dispersion with particles of test item visible dispersed throughout.

Results with reference substance (positive control):

3-Hour EC50 value of 8.5 mg/L, 95% confidence limits 6.7 - 11 mg/L

Table 1            Oxygen Consumption Rates and Percentage Inhibition Values after 3 Hours Contact Time

Nominal Concentration (mg/L)		Initial O2 Reading (mg O2/L)	Measurement Period (minutes)	Final O2Reading (mg O2/L)	O2Consumption Rates (mg O2/L/min)	% Inhibition
Control	R1	5.5	5	2.6	0.58	-
	R2	5.9	6	2.5	0.57	-
Test item	10	5.5	5	2.6	0.58	[1]
	32	5.4	5	2.6	0.56	3
	100	5.3	5	2.4	0.58	[1]
	320	5.3	6	1.8	0.58	[1]
	1000	6.2	8	1.9	0.54	6
3,5-dichlorophenol	3.2	6.3	8	2.8	0.44	23
	10	7.3	10	4.7	0.26	55
	32	8.1	10	7.3	0.08	86

[Increase in respiration rate as compared to controls]

R₁– R₂= Replicates 1 to 2

Table 2            pH Values of the Test Preparations at the Start and End of the Exposure Period

Nominal Concentration (mg/L)		pH
		0 Hours	3 Hours
Control	R1	7.3	7.8
	R2	7.6	7.8
Test item	10	7.5	7.8
	32	7.5	7.8
	100	7.6	7.8
	320	7.5	7.8
	1000	7.7	8.1
3,5-dichlorophenol	3.2	7.4	8.0
	10	7.4	8.1
	32	7.5	8.2

 

Table 3            Observations on the Test Preparations Throughout the Test Period

Nominal Concentration (mg/L)		Observations on Test Preparations
		0 Hours	30 Minutes Contact Time	3 Hours Contact Time
Control	R1	Dark brown dispersion	Dark brown dispersion	Dark brown dispersion
	R2	Dark brown dispersion	Dark brown dispersion	Dark brown dispersion
Test item	10	Extremely pale brown dispersion, brown particles of test item dispersed throughout[1]	Dark brown dispersion, few particles of test item visible dispersed throughout	Dark brown dispersion, few particles of test item visible dispersed throughout
	32	Pale brown dispersion, brown particles of test item dispersed throughout*	Dark brown dispersion, few particles of test item visible dispersed throughout	Dark brown dispersion, few particles of test item visible dispersed throughout
	100	Brown dispersion, brown particles of test item dispersed throughout*	Dark brown dispersion, particles of test item visible dispersed throughout	Dark brown dispersion, particles of test item visible dispersed throughout
	320	Dark brown dispersion, brown particles of test item dispersed throughout*	Dark brown dispersion, particles of test item visible dispersed throughout	Dark brown dispersion, particles of test item visible dispersed throughout
	1000	Very dark brown dispersion, brown particles of test item dispersed throughout*	Very dark brown dispersion, particles of test item visible dispersed throughout	Very dark brown dispersion, particles of test item visible dispersed throughout
3,5-dichlorophenol	3.2	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible
	10	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible
	32	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible	Dark brown dispersion, no undissolved reference item visible

R₁– R₂= Replicates 1 to 2

[1]Observations made prior to the addition of synthetic sewage and activated sewage sludge

R₁– R₂= Replicates 1 to 2

Validity criteria fulfilled:

yes

Conclusions:

The EC50 was greater than 1000 mg/L.

Executive summary:

The toxicity of the test material to microorganisms was assessed according to OECD Test Guideline 209 and EU Method C.11 and in compliance with GLP.

Under the conditions of the study the EC50 was greater than 1 000 mg/L.

Description of key information

The EC50  was greater than 1 000 mg/L.  

Additional information

The toxicity of the test material to microorganisms was assessed according to OECD Test Guideline 209 and EU Method C.11 and in compliance with GLP.

Under the conditions of the study the EC50 was greater than 1 000 mg/L.