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Toxicity to microorganisms

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Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
The study was well documented and meets generally accepted scientific principles, but was not conducted in compliance with GLP.
Qualifier:
equivalent or similar to guideline
Guideline:
other: ISO 10712
Principles of method if other than guideline:
Toxicity threshold test in Pseudomonas putida based on cell multiplication
GLP compliance:
not specified
Analytical monitoring:
not specified
Test organisms (species):
Pseudomonas putida
Details on inoculum:
- Laboratory culture: Stock cultures of Pseudomonas putida kept on nutrient medium in agar slant tubes. For onward culturing of the test strain, new stock cultures prepared at intervals of 1 week.

- Preparation of inoculum for exposure: Determine the extinction of the monochromatic radioation at 436 nm for a 10mm layer of the bacterial suspension by photoelectric measurement. On the bases of the values thus measured, adjust the final turbidity value of the bacterial suspension by means of sterile saline in such a way that the extinction value for a measuring sample that has been subject to onward dilution 1 + 9 with saline will correspond to the extinction value of a Formazin standard suspension TE/F/436nm = 10.
Test type:
other: Cell multiplication inhibition test
Water media type:
freshwater
Total exposure duration:
16 h
Remarks on exposure duration:
Incubation period
Test temperature:
25 degC
Details on test conditions:
TEST SYSTEM
- Test vessel: 300 ml Erlenmeyer flasks, stoppered with cotton-lined plastic cups

- Test vessel content: 80ml culture liquid, 5ml stock solution I, 5ml stock solution II, and 10ml each of bacterial suspension (or 10ml each of saline for the dilution series not inoculated)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Double distilled water

OTHER TEST CONDITIONS
- Adjustment of pH: Pollutant solution neutralised by addition of small amount acid or alkaline solution, in such a way that the volume added is kept as small as possible.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable)
- Concentration of the bacterial suspension measured turbidimetrically, and expressed as extinction of the primary light of monochromatic radiation at 436nm for a layer 10mm thick. The concentration at which the inhibitory action of a pollutant starts will be present in that step of a dilution series of the pollutant having an extinction value at the end of the test period that is ¿3% below the mean value of extinction for non-toxic dilutions of the test cultures.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: Four parallel dilution series, 1:2 to 1:2E+14 v/v. One series inoculated, one not.
Reference substance (positive control):
not required
Remarks:
not applicable
Duration:
16 h
Dose descriptor:
other: TT or EC3
Effect conc.:
62 mg/L
Basis for effect:
other: cell multiplication inhibition
Conclusions:
A toxicity threshold (TT) value for bacteria (Pseudomonas putida) of 62 mg/l was determined in a reliable study conducted according to generally accepted scientific principles.
Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
The study was well documented and meets generally accepted scientific principles, but was not conducted in compliance with GLP.
Qualifier:
no guideline followed
GLP compliance:
not specified
Analytical monitoring:
not specified
Test organisms (species):
other: other protozoa: Entosiphon sulcatum
Test type:
other: cell multiplication inhibition
Duration:
72 h
Dose descriptor:
other: TT or EC3
Effect conc.:
75 mg/L
Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
documentation insufficient for assessment
Remarks:
This information was obtained from the public IUCLID 2000 CD-ROM. Further review of the original source, to reassess the reliability, would not alter the overall conclusions concerning this endpoint.
Principles of method if other than guideline:
Method: other: not specified
GLP compliance:
not specified
Analytical monitoring:
not specified
Test organisms (species):
Tetrahymena pyriformis
Details on test conditions:
Type: aquatic
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
300.4 mg/L
Endpoint:
toxicity to microorganisms, other
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Remarks:
The result is sourced from secondary literature (Handbook). The original reference was not available for review and no further information is available.
Test organisms (species):
Uronema parduzci
Dose descriptor:
EC0
Effect conc.:
93 mg/L

Description of key information

Inhibition of WWTP microorganisms (hexan-1-ol): no significant inhibitory effects on respiration of activated sludges or specific microbial strains relevant to WWTP, at or above the limit of solubility (based on inhibition tests and lack of toxicity in ready biodegradability test).

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
62 mg/L

Additional information

In accordance with Column 2 of REACH Annex VIII, the activated sludge respiration inhibition study (required in Section 9.1.4 of REACH) does not need to be conducted as the substance is readily biodegradable and the applied test concentrations are in the range that can be expected in the influent to a sewage treatment plant.

Among the available existing data there is one reliability 2 study available, for the species Pseudomonas putida. A 16-h toxicity threshold of 62 mg/L was determined. This is supported by consistent data for other micro-organism species. The available data are taken into account in environmental modelling. 

This is supported by results of non-assignable reliability studies with longer chain-lengths within the C6 -24 Alcohols Category, indicating EC50in the hundreds of mg/l (in some cases, up to and exceeding the limit of solubility) for respiration of a mixed microbial culture.

Discussion of trends in the Category of C6-24 linear and essentially-linear aliphatic alcohols: 

A number of studies of the toxicity of LCAAs to single species of micro-organisms have been identified. A range of results are shown in Table 6.1 of the ECOTOXICITY Alcohols C6-24 Category report, for test species relevant to WWTP (studies on various other organisms are also available). 

 

The ready biodegradation studies (Federle, 2009 and Flach, 2012, discussed in the environmental fate section) gives evidence that rapid biodegradation by aquatic activated sludge microorganisms is seen, even in the presence of an excess of test substance (for the longer chain length category members tested).

 

At or above the limit of solubility, the data set shows that the alcohols in the chain length range C6-24 category have no significant inhibitory effects on respiration of activated sludges or specific microbial strains relevant to WWTP. There is limited evidence of inhibition of growth in some specific microbial strains but ready biodegradation evidence suggests this is not significant for mixed populations.

 

The reliability of some of these data individually is not high, but it presents a consistent weight of evidence.

 

In general it can be said that the micro-organisms examined in these tests were less susceptible to the LCAAs than fish, invertebrates and algae and that in general, IC50 values for respiration effects are at or significantly above the limit of water solubility.

 

WWTP microorganisms have been demonstrated to be capable of synthesising significant concentrations of aliphatic alcohols (e.g. Mudge et al., 2008).

Table: Micro-organism toxicity for species of potential relevance to WWTP systems

CAS

Chemical Name

Comment

Water solubility (mg/L)

Species

Endpoint

Result (mg/L)

Effect

Reliability

Reference

111-27-3

Hexan-1-ol

 

6320

Pseudomonas putida

16-hr Toxicity Threshold (TT)

62

Cell multiplication (similar to ISO 10712)

2

Bringmann and Kuhn, 1980

111-27-3

Hexan-1-ol

 

6320

Uronema parduczi

EC0

93

Not stated

4

Verschueren, 1996

111-27-3

Hexan-1-ol

 

6320

P. putida

30 min EC10

3000

Cell multiplication (DIN 38412 Part 8)

4

Henkel unpublished

111-27-3

Hexan-1-ol

 

6320

P. putida

16-hr EC10

10,000

Respiration (DIN 38412 Part 27)

4

unreferenced; IUCLID 2000 CD-ROM

111-70-6

Heptan-1-ol

Supporting

1410

P. putida

16-hr Toxicity Threshold (TT)

67

Cell multiplication (similar to ISO 10712)

2

Bringmann and Kuhn, 1980

111-70-6

Heptan-1-ol

Supporting

1410

U. parduczi

EC0

17

Not stated

4

Verschueren, 1996

111-87-5

Octan-1-ol

 

551

P. putida

16-hr Toxicity Threshold (TT)

>50

Cell multiplication (similar to ISO 10712)

2

Bringmann and Kuhn, 1980

111-87-5

Octan-1-ol

 

551

Chilomonas paramecium

48 h EC5

>20

Cell multiplication

4

Verschueren, 1996

111-87-5

Octan-1-ol

 

551

U. parduczi

20 h TGK 5%

23

Cell multiplication

4

Bringmann and Kuhn, 1980

111-87-5

Octan-1-ol

 

551

Clostridium acetobutylicum(anaerobic)

6-h EC50

130.3

Growth

4

Izardet al., 1989

111-87-5

Octan-1-ol

 

551

 Activated sludge

3 h EC50

350

Respiration (OECD 209)

4

Tanget al., 1990

111-87-5

Octan-1-ol

 

551

 Activated sludge

EC50

325.6

Heat flux

4

Beaubienet al., 1985

111-87-5

Octan-1-ol

 

551

 Activated sludge

24h EC50

200

Respiration (other methodology)

4

Tanget al., 1990

111-87-5

Octan-1-ol

 

551

 P. putida

30 min EC10

10,000

Respiration (DIN 38412 Part 27)

4

Bringmann and Kuhn, 1980

Henkel (undated)

111-87-5

Octan-1-ol

 

551

 Mixed culture

75 min EC50

625

Respiration

4

Vaishnavet al., 1986

112-30-1

Decan-1-ol

 

39.5

P. putida

30-min. EC0

10,000

Respiration (DIN 38412 Part 27)

2

Henkel, 1999a

112-30-1

Decan-1-ol

 

39.5

Mixed microbial culture

75 min EC50

443

Respiration

4

Vaishnavet al., 1985

112-30-1

Decan-1-ol

 

39.5

 P. putida

30 min EC0

10,000

Respiration

4

Henkel, unpublished

112-53-8

Dodecan-1-ol

 

2.07

 P. putida

30-min. EC0

>10,000

Respiration (DIN 38412Part27)

1

Henkel, 1994b

80206-82-2

C12-14 Alcohols

Supporting

 

P. putida

30 min EC0

>10,000

Respiration (DIN 38412Part27)

4

Henkel unpublished

80206-82-2

C12-14 Alcohols

Supporting

 

P. putida

30 min EC0

10,000

Respiration (DIN 38412Part27)

4

Henkel unpublished

112-72-1

Tetradecan-1-ol

 

0.191

P. putida

30 min EC50

>10,000

Respiration (DIN 38412Part27)

4

Henkel unpublished

68002-94-8

C16-18 and C18 Unsturated

Supporting

 

P. putida 

30-min. EC10

>10,000

Respiration (DIN 38412Part27)

2

Henkel, 1988a

112-92-5

Octadecan-1-ol

 

0.0011

P. putida

30-min.EC0

>10,000

Respiration (DIN 38412Part27)

1

Henkel, 1994c

References:

Bringmann, G. and Kuhn, R. 1980. Comparison of the toxicity thresholds of water pollutants to bacteria, algae, protozoa in the cell multiplication inhibition test.Water Research 14:231-241.

Verschueren, K. (ed.). 1996. Handbook of Environmental Data on Organic Chemicals. 3rd ed. New York: John Wiley & Sons, Inc.

Izardet al., 1989:  Appl. Microbiol. Biotechnol. 31, 179-183 (1989).

Tang, N.H.et al., J. Environ.Eng. (New York) 116, 1076-1084 (1990)

Beaubien, A.et al., Water Res. 19, 747-755 (1985)Beilstein On-Line database.

Vaishnav, D.D., Toxicity Assessment 1, 227-240 (1986)

Henkel KGaA.1999a. 1-Decanol: Acute toxicity bacteria. Unpublished results. Biological Research and Product Safety/Ecology. Test Substance Registration No. 7200.

Vaishnav, D.D. and Lopas, D.M. Dev. Industr. Microbiol. 26 (1985), 557-565

Henkel KGaA.1994b. Report-Nummer R 9400236. A. Kirch. Marz 1994.

Henkel KGaA. 1988a.Biological Research and Product Safety/Ecology. Test substance registration No. 6802. Bacterial toxicity.

Henkel KGaA. 1994c.1-Octadecanol: Sauerstoffzehrungshemmtest mit Bakterian. Abschlubericht. Report-Nummer R 9400049. A. Kirch. Januar 1994.

Henkel KGaA, unpublished data (1), Archive-No. 7198 (no further details available); cited in IUCLID 2000 CD-ROM (source documentation not available)

Henkel KGaA, unpublished data (2); cited in IUCLID 2000 CD-ROM (source documentation not available)

Henkel KGaA, unpublished data (3), Final Report 882633 (no further details available); cited in IUCLID 2000 CD-ROM (source documentation not available)

Henkel KGaA, unpublished data (4), Registry No. 6801 (no further details available); cited in IUCLID 2000 CD-ROM (source documentation not available)

Henkel KGaA, unpublished data (5), File 427/1 (no further details available); cited in IUCLID 2000 CD-ROM (source documentation not available)

Henkel KGaA, unpublished data (6), Test No. 9300287 31; Final Report 9500603 (no further details available); cited in IUCLID 2000 CD-ROM (source documentation not available)