Magnesium chloride

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: study generated according to valid and internationally accepted testing guidelines and performed according to GLP

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

certified laboratory

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Two stock solutions, S1 (100 mg/L) and S2 (10 mg/L) were prepared and diluted stepwise to achieve the required test concentrations 1, 01, and 0.01 mg/L (D1 – D3)

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Activated sludge from the municipal wastewater treatment plant of Pforzheim, Germany, was used as the microbial inoculum for the test. This plant is pre-dominantly treating domestic sewage.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Post exposure observation period:

Not approriate

Hardness:

no data

Test temperature:

20.0 – 21.1 °C (during O2 measurement)

pH:

8.01 – 8.10

Dissolved oxygen:

The rate of respiration for each flask was measured over the linear portion of the oxygen consumption trace.

Salinity:

No data

Nominal and measured concentrations:

Nominal concentrations :
- A range-finding test with concentrations from 0.01 to 100 mg/L (nominal), in dilution steps of 10-1 was performed.
- Subse-quently, a limit test with six times 100 mg/L (nominal) was performed,
- and finally a confirmatory test with two untreated controls, test item rates of 12, 35, 100, 300 and 900 mg/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 1 litre glass beakers
- Type (delete if not applicable): open
- Aeration: the test was aerated.
- No. of organisms per vessel:
- No. of vessels per concentration (replicates): 6 replicates
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test water used for the definitive test was laboratory tap water dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex water softener).

OTHER TEST CONDITIONS
- Adjustment of pH: pH was 8.01 – 8.10 and was not adjusted
- Light intensity: Normal laboratory lighting

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
The rate of respiration for each flask was measured over the linear portion of the oxygen consumption trace

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
A range-finding test with concentrations from 0.01 to 100 mg/L, in dilution steps of 10-1 was performed. Subse-quently, a limit test with six times 100 mg/L was performed, and finally a confirmatory test with two untreated controls, test item rates of 12, 35, 100, 300 and 900 mg/L and three concentration of 3,5-Dichlorophenol as a toxic reference item to demonstrate the sensitivity of the test system.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol (Sigma-Aldrich Batch No. 04621CJ)

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 900 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

No effects were found in the range-finding test. The main test (limit test), therefore, was performed in a limit test design with six replicates of 100 mg/L. Since slight inhibitory effects were found in the test limit at 100 mg/L, a confirmatory test from 12 mg/L to 900 mg/L was prepared to show whether the inhibitions observed in the limit test were real toxic effects. The results of this confirmatory test are presented below: These results confirms that no-effect were observed between 12 and 900 mg/L, and thus confirmed that Magnesium Chloride can be regarded as non-toxic to activated sludge organisms (EC50 > 900 mg/L, NOEC = 900 mg/L).

Results with reference substance (positive control):

The EC50 value for DCP was in the range between 5 and 30 mg/L. (13.65 mg/L after 30 min and 7.58 mg/L after 3 h).
Therefore, the test fulfils the criteria for validity.

Confirmatory test was performed in order to examine if the results of the limit test at 100 mg were real inhibitory effects. The results of the confirmatory test are summarised in the following Tables.

Table 1. Results of the confirmatory test, respiration rates

Magnesium Chloride [mg/L]	Respiration rates [mg O2/L · h]
	30 min	3 h
0	21.60	17.49
12	21.92	17.31
35	21.90	17.43
100	23.12	18.54
300	22.40	17.94
900	23.43	18.29
DCP 5 mg/L	17.87	11.22
DCP 15 mg/L	9.87	5.40
DCP 30 mg/L	5.51	2.97
0	22.11	18.53

Table 2 : Results of the confirmatory test, percent inhibition

Magnesium Chloride [mg/L]	% inhibition
	30 min	3 h
Control 1	1.17*	2.89*
12	-0.30	3.89
35	-0.21	3.22
100	-5.79	-2.94
300	-2.49	0.39
900	-7.21	-1.55
DCP 5 mg/L	18.23	37.70
DCP 15 mg/L	54.84	70.02
DCP 30 mg/L	74.79	83.51
Control 2	-1.17*	-2.89*

*Percent deviation from the mean values of both controls

Validity criteria fulfilled:

yes

Remarks:

All validity criteria of OECD Guideline for Testing of Chemicals No. 209 were fulfilled

Conclusions:

The effect of the test material on the respiration of activated sewage sludge micro-organisms gave a 3-Hour EC50 of greater than 900 mg/L, and the No Observed Effect Concentration (NOEC) after 3 hours exposure was 900 mg/L.

Executive summary:

The respiration rate of an activated sludge, fed with a standard amount of synthetic wastewater feed, was measured after 30 minutes and 3 hours of contact in presence of the test item. The inhibitory effect of Magnesium Chloride hexahydrate of particular concentrations was expressed as a percentage of the mean respiration rate of two controls.

Activated sludge from the municipal wastewater treatment plant of), was used as microbial inoculum for the test. This plant predominantly is treating domestic sewage.

The sludge was adjusted to a content of 1.6 g/L of mixed liquid suspended solidsand was exposed to the test item under continuous aeration. After stopping the aeration, the O₂consumption was measured for approx. 10 minutes. The slope of the O₂consumption straight line is an indication for toxic effects on respiration activity of microorganisms.

Three series of tests were successively performed:

1. Range-finding tests :

First, a range-finding test with concentrations from 0.01 to 100 mg/L, in dilution steps of 10^-1was performed in order to determine the range of concentrations for the main test.

2. Main test (Limit test) :

Subsequently, a limit test with six replicates contaminated at 100 mg/L was performed. Slight inhibitory effects were found at any replicate.

3. Confirmatory test :

Finally a further test (confirmatory test) from 12 mg/L to 900 mg/L (12, 35, 100, 300 and 900 mg/L) was performed to show whether the inhibitions observed in the limit test were real toxic effects. No effects were found in this confirmatory test.

Hence, based on the results of this last confirmatory test an EC50 may safely be expected above 900 mg/L, i.e. EC50 > 900 mg/L. A NOEC of 900 mg/L can thus be determined for Magnesium chloride hexahydrate.

 

Description of key information

An Activated Sludge, Respiration Inhibition Test (ASRIT) has been performed on Magnesium Chloride hexahydrate in accordance with GLP and OECD 209.

No short-term toxicity to microorganisms was observed at the concentration tested (12 -900 mg test item/L).

Hence, the effect of the test material on the respiration of activated sewage sludge micro-organisms gave a 3-Hour EC50 of greater than 900 mg of MgCL2 6H20/L. The No Observed Effect Concentration (NOEC) after 3 hours exposure was 900 mg MgCl2 6H2O/L corresponding to 421 mg MgCl2/L.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

421 mg/L

Additional information

The toxicity toward the microorganisms of MgCl2 was investigated was investigated in one GLP-compliant study performed in accordance with standard methods, without deviations. The study is considered as reliable (Klimisch 1) and is selected as a key study for the endpoint.

A further supporting study, reported in the IUCLID Dataset (2000) reports a 30min EC50s of 9266 mg of Mg/L for the respiration of activated sewage sludge micro-organisms.

A third study performed on Vibrio fischeri is considered not relevant for Sewage Treatment Plant as the bacterium is a saltwater species.