Methenamine

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to microorganisms, other

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1995

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: non GLP, no guideline

Qualifier:

according to guideline

Guideline:

DIN 38412-8 (Pseudomonas Zellvermehrungshemmtest)

Deviations:

yes

Remarks:

additional time points 60 and 90 min

Principles of method if other than guideline:

Measurement of luminescence

GLP compliance:

not specified

Analytical monitoring:

no

Details on sampling:

no sampling

Vehicle:

no

Details on test solutions:

no details given

Test organisms (species):

Vibrio fisheri

Details on inoculum:

The marine, luminescent bacteria strain Vibrio fischeri NRRL-B-11177 (formerly Photobacterium phosphoreum; DSM 7151) was used for luminescence inhibition test system. In order to ensure optimal salt conditions for the bacteria, the samples must have a salt content of 2%, which was achieved by adding 2% NaCl in solid form.

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

30 min

Remarks on exposure duration:

60 and 90 min also tested

Post exposure observation period:

no post exposure period

Hardness:

not indicated

Test temperature:

15°C

pH:

not indicated

Dissolved oxygen:

not indicated

Salinity:

2 %

Nominal and measured concentrations:

5000 mg/L (nominal)

Details on test conditions:

The LUMIStox measuring unit from Dr. Lange GmbH (Berlin, Germany) was used to determine the ecotoxicological potential of Hexamethylenetetramine. The EC50 values were determined. The sample sequence corresponds to DIN 38412 L34 (L341), which ranges from 1:2 to 1:32-dilution of the compound in the test system. All dilution steps were made in duplicate. The change of intensity of bacterial luminescence in the absence (controls) and in the presence (samples) of Hexamethylenetetramine after different incubation times (30 min according to DIN 38412 and after 60 and 90 min) were recorded. The controls (with 2% NaCl only) were measured for calculating the correction factor, which was necessary to consider the "normal" decrease of luminescence (without any toxic effect) per time. The corrected values were used for calculating the percentage of inhibition of luminescence per incubation period of each sample. The bioluminescence of the bacteria is measured by a temperature-controlled photomultipier at 15°C.

Reference substance (positive control):

yes

Remarks:

other substances with positive effects were measured in parallel.

Duration:

30 min

Dose descriptor:

EC50

Effect conc.:

> 5 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: luminescence

Key result

Duration:

60 min

Dose descriptor:

EC50

Effect conc.:

> 5 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: luminescence

Key result

Duration:

90 min

Dose descriptor:

EC50

Effect conc.:

> 5 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: luminescence

Details on results:

no details given

Results with reference substance (positive control):

Other substances with positive effects were measured in parallel.

Reported statistics and error estimates:

not indicated

Validity criteria fulfilled:

yes

Conclusions:

The LC50 of methenamine in Vibiro fischeri was determined to be > 5000 mg/L.

Executive summary:

In a 30, 60 and 90 min toxicity study, cultures of Vibrio fischeri were exposed to methenamine at a nominal limit-concentrations of 5000 mg/L under static conditions. The NOEC and EC50 values based are based on luminescence. 5000 mg/l, the highest concentration tested, did not reveal any effects for a test duration of up to 90 min. Thus the EC50 and the NOEC values were > 5000 mg/L.

This toxicity study is classified as acceptable for the risk assessment for STPs.

Results Synopsis

Test Organism: Vibrio fischeri

Test Type: Static

90 min EC50: > 5000 mg/L

90 min NOEC: >= 5000 mg/L

Reference 2

Endpoint:

activated sludge nitrification inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1977

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: non GLP, no guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Measurement of inhibition of nitrification.

GLP compliance:

not specified

Analytical monitoring:

no

Details on sampling:

no sampling

Vehicle:

no

Details on test solutions:

not indicated

Test organisms (species):

other: Nitrosomas sp. and Nitrobacter sp.

Details on inoculum:

autotrophic nitrifying bacteria

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

2 h

Post exposure observation period:

no post exposure period

Hardness:

not indicated

Test temperature:

not indicated

pH:

8.1

Dissolved oxygen:

not indicated

Salinity:

not indicated

Nominal and measured concentrations:

nominal: 100 mg/L

Details on test conditions:

Throughout the experiments a stock solution of autotrophic nitrifying bacteria was maintained under continuous culture conditions. Organisms were removed from this culture and used in a standard assay to determine the effects of the Hexamethylenetetramine by comparing the rate of nitrate production in the presence of the test compound to the rate in its absence. All determinations were made in duplicate, and Hexamethylenetetramine was considered to be inhibitory if it reduced the rate by more than 10 %.

Reference substance (positive control):

yes

Remarks:

other substances with positive response were tested in parallel.

Key result

Duration:

2 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of nitrification rate

Key result

Duration:

2 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of nitrification rate

Details on results:

100 mg/l, the highest concentration tested, did not reveal any effects for a test duration of 2 hours.

Results with reference substance (positive control):

Other substances with positive response were tested in parallel.

Reported statistics and error estimates:

not indicated

Validity criteria fulfilled:

yes

Conclusions:

The LC50 of methenamine in nitrifying bacteria (Nitrosomas sp. and Nitrobacter sp.) was determined to be > 100 mg/L.

Executive summary:

In a 2 hour acute toxicity study, cultures of nitrifying bacteria (Nitrosomas sp. and Nitrobacter sp.) were exposed to methenamine at a nominal limit-concentrations of 100 mg/L under static conditions. The NOEC and EC50 values based are based on nitrification (production of nitrate). 100 mg/l, the highest concentration tested, did not reveal any effects for a test duration of 2 hours. Thus the EC50 and the NOEC values were > 100 mg/L.

This toxicity study is classified as acceptable for the risk assessment.

Results Synopsis

Test Organism: nitrifying bacteria (Nitrosomas sp. and Nitrobacter sp.)

Test Type: Static

2 hr EC50: > 100 mg/L

2 hr NOEC: >= 100 mg/L

Description of key information

The LC50 of methenamine in nitrifying bacteria (Nitrosomas sp. and Nitrobacter sp.) was determined to be > 100 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:

100 mg/L

EC10 or NOEC for microorganisms:

100 mg/L

Additional information

Key studies:

- Freshwater: In a 2 hour acute toxicity study (Hockenbury, 1977), cultures of nitrifying bacteria (Nitrosomas sp. and Nitrobacter sp.) were exposed to methenamine at a nominal limit-concentration of 100 mg/L under static conditions. The NOEC and EC50 values are based on nitrification (production of nitrate). 100 mg/L, the highest concentration tested, did not reveal any effects for a test duration of 2 hours. Thus the EC50 and the NOEC values were > 100 mg/L.

This toxicity study is classified as acceptable for the risk assessment.

- Saltwater: In a 30, 60 and 90 min toxicity study (Drzyzga, 1995), cultures of Vibrio fischeri were exposed to methenamine at a nominal limit-concentrations of 5000 mg/L under static conditions. The NOEC and EC50 values based are based on luminescence. 5000 mg/l, the highest concentration tested, did not reveal any effects for a test duration of up to 90 min. Thus the EC50 and the NOEC values were > 5000 mg/L.

This toxicity study is classified as acceptable for the risk assessment.

 

Supporting study:

- In a non standard assay (Greenwood, 1981) the inhibitory effect of bacteria growth of methenamine was examined. Different strains of Escherichia coli and Klebsiella aerogenes showed MAC-values (minimum antibacterial concentration = lowest concentration which caused partial inhibition of growth) between 32 and 125 mg/L and MIC-values (minimum inhibitory concentration = lowest concentration which inhibited growth overnight) between 250 and 500 mg/L. No NOECs or EC50 values are given or can be calculated, based on the data presented. Thus this study cannot be used for risk assessment purposes.

 

Based on the available data the NOEC > 100 mg/L observed in Nitrosomas sp. and Nitrobacter sp. can be considered as a conservative reference figure for the risk assessment of STPs.