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Ecotoxicological information

Toxicity to microorganisms

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Reference
Endpoint:
toxicity to microorganisms, other
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Remarks:
Experimental data of test chemical
Justification for type of information:
Experimental data for test chemical is from handbook or collection of data
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
WoE report is prepared based on long term toxicity to fish study:
1 and 2nd study
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
not specified
Details on test solutions:
2 study: Details on test solutions
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Determination of MIC was by the broth dilution method
Test organisms (species):
other: 1: Colletotrichum musae DAR 24962, 2: Bacillus subtilis
Details on inoculum:
1: The surface-plated cultures of the decay fungi in plastic Petri dishes were sub-cultured by streaking the spores onto the new potato dextrose agar (PDA) media. The plated cultures were then incubated for 7 days at 25°C.
The spores of 7-day-old cultures of decay fungi dislodged by sterile distilled water to which 0.1 mL/L of Tween 80 had been added. The spores were then filtered with sterile Sinta Glass No. 1 (Gallenkamp, London) to remove debris such as mycelia and condensed agar fragments, and the aliquot was diluted to a concentration of 10(5) fungal spores/mL suspensions. The fungal spore suspensions, 0.1 mL each, were then dispensed into Petri dishes (9-cm diam.) containing agar medium (PDA). The Petri dishes were then incubated for 3 days for the fungal cultures at 25 °C, to allow the spores to grow.

2nd study: Details on test organisms:
2-day-old cultures of the microorganisms were used in the test
Test type:
other: 1: static, 2: broth dilution method
Water media type:
freshwater
Total exposure duration:
10 d
Post exposure observation period:
1. Agar disks of microorganism which failed to grow due to MIC were transferred onto new agar media free from test chemical and incubated for further 5 days at 25°C.
2: 2-5 days
Test temperature:
25°C
Nominal and measured concentrations:
1. 4.65 mmol/dish (equivalent to 894.195 mg/L/dish)
Details on test conditions:
1. Agar plugs (5.5-mm diam.) were picked up from the 3-day-old cultures of decay fungi using the bottom end of a sterilized Pasteur pipet and then transferred ontothe centers of new PDA media, in 9-cm plastic Petri dishes. The Petri dishes were then inverted and 7-cm Whatman No. 1 filter papers were attached onto the inner surface of their lids.
Ethanol, the first tested volatile in this experiment, was impregnated into the filter paper with varying volumes from 0.1 to 1.0 mL/dish in the 4°C room. Immediately after the impregnation, the Petri dishes were sealed by wrapping them with plastic film and incubated for 10 days at 25 °C. Experiments were repeated two timeswith four replications for each experiment. The minimum concentration of ethanol (expressed as mmol/dish) required to give complete control or the minimum inhibitory concentration (MIC) for each microorganism was determined.
The MIC of ethanol for each target decay microorganism was used as the initial level to identify the MIC of other tested volatiles. If the MIC level of ethanol used for other volatiles failed to stop the growth of pathogen, the level was increased until the MIC was found. However, if the volume of 1.5 mL/dish still failed to stop the growth of pathogen, the compound was considered ineffective as a vapor to stop the growth of pathogens. When the tested compounds had the same effect as the MIC of ethanol, the concentration was decreased until the MIC of the compound for each microorganism was determined. All the unit concentrations of MIC were then expressed as mmol/dish.
Reference substance (positive control):
yes
Remarks:
1: Ethanol
Key result
Duration:
10 d
Dose descriptor:
other: minimum inhibitory concentration (MIC)
Effect conc.:
894.195 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Remarks on result:
other: 1st study
Key result
Duration:
5 d
Dose descriptor:
other: MIC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Remarks on result:
other: 2ND STUDY
Details on results:
1. Test chemical exhibited fungistatic activity
Results with reference substance (positive control):
1. Ethanol was considered germistatic against C. musae with MIC of 16.59 mmol/dish
Validity criteria fulfilled:
not specified
Conclusions:
1. The Minimum Inhibitory Concentration (MIC) value of test chemical on the fungi Colletotrichum musae DAR 24962 was determine to be 894.195 mg/L.
2. Based on the antimicrobial effect of test chemical on the growth of Bacillus subtilis after the exposure of chemical for 2-5 days, the MIC was observed at 100 mg/l.
Thus based on the above studies, MIC ranges from 100 mg/l to 894.195 mg/l after the exposure of microorganisms with the test chemical for 2-10 days.
Executive summary:

Based on the various experimental data for the test chemical study have been reviewed to determine the toxic nature of test chemical on the growth of microorganisms. The studies are as mentioned below:  

The effects of test chemical on the growth of Colletotrichum musae on agar medium were evaluated. Test was performed on the agar medium. Agar plugs (5.5-mm diam.) were picked up from the 3-day-old cultures of decay fungi using the bottom end of a sterilized Pasteur pipet and then transferred onto the centers of new PDA media, in 9-cm plastic Petri dishes. The Petri dishes were then inverted and 7-cm Whatman No. 1 filter papers were attached onto the inner surface of their lids. Ethanol, the first tested volatile in this experiment, was impregnated into the filter paper with varying volumes from 0.1 to 1.0 mL/dish in the 4°C room. Immediately after the impregnation, the Petri dishes were sealed by wrapping them with plastic film and incubated for 10 days at 25 °C. Experiments were repeated two times with four replications for each experiment. The minimum concentration of ethanol (expressed as mmol/dish) required to give complete control or the minimum inhibitory concentration (MIC) for each microorganism was determined. The MIC of ethanol for target decay microorganism was used as the initial level to identify the MIC of other tested volatiles. If the MIC level of ethanol used for other volatiles failed to stop the growth of pathogen, the level was increased until the MIC was found. However, if the volume of 1.5 mL/dish still failed to stop the growth of pathogen, the compound was considered ineffective as a vapor to stop the growth of pathogens. When the tested compounds had the same effect as the MIC of ethanol, the concentration was decreased until the MIC of the compound for each microorganism was determined. All the unit concentrations of MIC were then expressed as mmol/dish. After the incubation of 10 days, the Minimum Inhibitory Concentration (MIC) value of test chemical on the fungi, Colletotrichum musae DAR 24962 was determine to be 894.195 mg/L.

 

First study was supported by the second experimental study. Aim of this study was to evaluate the effect of test chemical on the growth of Bacillus subtilis and other fungi. The antimicrobial activity of test compounds against various bacteria and fungi was examined by the broth dilution method. Solution of the test compound was added to 2-day-old cultures of the microorganisms. After 2-5 days of incubation, growth of the microorganisms was checked. Minimal inhibitory concentrations (MICs) were measured by two fold serial broth dilution. Based on the antimicrobial effect of test chemical on the growth of Bacillus subtilis after the exposure of chemical for 2-5 days, the MIC was observed at 100 mg/l.

 

Thus based on the above studies, MIC ranges from 100 mg/l to 894.195 mg/l after the exposure of microorganisms with the test chemical for 2-10 days.

Description of key information

1. The Minimum Inhibitory Concentration (MIC) value of test chemical on the fungi  Colletotrichum musae DAR 24962 was determine to be 894.195 mg/L.

2. Based on the antimicrobial effect of test chemical on the growth of Bacillus subtilis after the exposure of chemical for 2-5 days, the MIC was observed at 100 mg/l.

Thus based on the above studies, MIC ranges from 100 mg/l to 894.195 mg/l after the exposure of microorganisms with the test chemical for 2-10 days.

Key value for chemical safety assessment

EC50 for microorganisms:
100 mg/L

Additional information

Based on the various experimental data for the test chemical and structually and functionally similar read across chemicals, study have been reviewed to determine the toxic nature of test chemical on the growth of microorganisms. The studies are as mentioned below:  

The effects of test chemical on the growth of Colletotrichum musae on agar medium were evaluated. Test was performed on the agar medium. Agar plugs (5.5-mm diam.) were picked up from the 3-day-old cultures of decay fungi using the bottom end of a sterilized Pasteur pipet and then transferred onto the centers of new PDA media, in 9-cm plastic Petri dishes. The Petri dishes were then inverted and 7-cm Whatman No. 1 filter papers were attached onto the inner surface of their lids. Ethanol, the first tested volatile in this experiment, was impregnated into the filter paper with varying volumes from 0.1 to 1.0 mL/dish in the 4°C room. Immediately after the impregnation, the Petri dishes were sealed by wrapping them with plastic film and incubated for 10 days at 25 °C. Experiments were repeated two times with four replications for each experiment. The minimum concentration of ethanol (expressed as mmol/dish) required to give complete control or the minimum inhibitory concentration (MIC) for each microorganism was determined. The MIC of ethanol for target decay microorganism was used as the initial level to identify the MIC of other tested volatiles. If the MIC level of ethanol used for other volatiles failed to stop the growth of pathogen, the level was increased until the MIC was found. However, if the volume of 1.5 mL/dish still failed to stop the growth of pathogen, the compound was considered ineffective as a vapor to stop the growth of pathogens. When the tested compounds had the same effect as the MIC of ethanol, the concentration was decreased until the MIC of the compound for each microorganism was determined. All the unit concentrations of MIC were then expressed as mmol/dish. After the incubation of 10 days, the Minimum Inhibitory Concentration (MIC) value of test chemical on the fungi, Colletotrichum musae DAR 24962 was determine to be 894.195 mg/L.

 

First study was supported by the second experimental study. Aim of this study was to evaluate the effect of test chemical on the growth of Bacillus subtilis and other fungi. The antimicrobial activity of test compounds against various bacteria and fungi was examined by the broth dilution method. Solution of the test compound was added to 2-day-old cultures of the microorganisms. After 2-5 days of incubation, growth of the microorganisms was checked. Minimal inhibitory concentrations (MICs) were measured by two fold serial broth dilution. Based on the antimicrobial effect of test chemical on the growth of Bacillus subtilis after the exposure of chemical for 2-5 days, the MIC was observed at 100 mg/l.

 

Thus based on the above studies, MIC ranges from 100 mg/l to 894.195 mg/l after the exposure of microorganisms with the test chemical for 2-10 days.