Anthracene oil

Toxicity to soil microorganisms

Administrative data

Endpoint:

toxicity to soil microorganisms

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

Data source

Reference

Reference Type:

publication

Title:

Soil microbial toxicity of eight polycyclic aromatic compounds: Effects on nitrification, the genetic diversity of bacteria, and the total number of protozoans

Author:

Sverdrup LE, Ekelund F, Krogh PH, Nielsen N, Lohnson K

Year:

2002

Bibliographic source:

Environ Toxicol Chem 21, 1644-1650

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

ISO 14238

Deviations:

yes

Remarks:

drying/dehumidification, use of solvent, addition of fresh inoculum derived from soil

Principles of method if other than guideline:

Comparative study including four aromatic and four heteroaromatic compounds. In addition to nitrification, other parameters were analysed (the genetic diversity of bacteria, total number of protozoans).

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

Phenanthrene

EC Number:

201-581-5

EC Name:

Phenanthrene

Cas Number:

85-01-8

Molecular formula:

C14H10

IUPAC Name:

phenanthrene

Test material form:

solid

Details on test material:

- Substance type: organic
- for additional information see study record

Specific details on test material used for the study:

TEST MATERIAL
- Name of test material (as cited in study report): phenanthrene
- Source of test material: Sigma-Aldrich, St. Louis, MO, USA
- Analytical purity: > 96 %

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 10 mg/kg soil dw (low conc.) and 1000 mg/kg soil dw (high conc.)
- Sampling time: start of the experiment
- Sample size: 1000 mg soil for low concentration and 100 mg soil for high concentration

Test substrate

Vehicle:

yes

Remarks:

acetone

Details on preparation and application of test substrate:

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method of preparation of test solutions: a stock solution in acetone was prepared that corresponded to the highest test dosage. For each test concentration, dilutions from the stock solution with acetone were made such that 4 mL of the resulting acetone solution contained the amount of test material required to produce the final test concentration after mxing with soil.
- Method of mixing into soil: 4 mL acetone solution for each of the treatments (equal volumes) was added to a 20 g soil dw in a glass vessel and thoroughly mixed into the soil resulting in the desired test concentration. The solvent was evaporated for 24 h.
- Controls: without and with solvent (soil samples were treated the same way as the test samples but using acetone without test substance)

VEHICLE
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution): not specified
- Evaporation of vehicle before use: yes

AMENDMENT OF SOIL
- Nitrogen source: horn meal (Solsikken, Lina, Denmark) at 1 ± 0.05 g/kg soil dw was added to each glass vessel (horn meal nitrogen content: 9.1 %, C/N ratio. 3.48 (mol/mol))
- Inoculum: 3.5 mL inoculum prepared from fresh Askov soil was added to each soil sample (glass container with 20 g soil dw) and thoroughly mixed. The Askov soil samples used in the test were pretreated (dried at 80 °C). Therefore inoculum from fresh Askov soil (stored at 5 °C for a maximum of 3 days) was added to the test samples.
preparation of inoculum according to a procedure described by Lindahl and Bakken (1995, FEMS Microbiol Ecol 16:135-142). In general, a slurry of 20 g fresh soil in 180 mL of deionised water was prepared by repeated mixing with a kitrchen blender for one minute followed by cooling on ice. The amount of water within the 3.5 mL inoculum added to the microcosms corresponded to 57 % of the water holding capacity of the soil. The population density of microorganisms added was estimated to be 2 % of that in the fresh material (100 % extraction efficiency assumed).

Test organisms

Test organisms (inoculum):

soil

Study design

Total exposure duration:

4 wk

Test conditions

Test temperature:

20 °C

Moisture:

57 ± 5 % of water holding capacity; gravimetric determination of water loss;
twice a week during incubation period water was added corresponding to water loss

Organic carbon content (% dry weight):

1.6

Nitrogen content (% dry weight):

0.443

Details on test conditions:

TEST SYSTEM
- Test container: cylinders (5 cm height, 3.5 cm diameter); sealed with plastic lids; upper lid perforated with five holes (made by a needle); at the start of the test, the inoculated soil samples (see above) were tranferred from the glass vessels to the test containers
- Amount of soil: 20 g
- No. of replicates per concentration: 3
- No. of replicates per control: 3
- No. of replicates per vehicle control: 3

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Soil test substrate: Danish agricultural soil
- Geographic location: Askov, Jutland, Denmark
- Soil texture (if natural soil): sandy loam
- Composition: coarse sand (0.2 – 2 mm) 38.4 %,
fine sand (0.063 – 0.2 mm) 23.6 %,
coarse silt (0.02 – 0.063 mm) 10.0 %,
fine silt (0.002 – 0.02 mm) 12.3 %,
clay (<0.002 mm) 13 %,
humus 2.8 %
- Organic carbon content: 1.6 %
- Soil pH: 6.2, no adjustment
- Density: 1.135 g/cm³ dry soil
- Maximum water holding capacity (in % dry weight): no data
- CEC (Cation exchange capacity): 8.14 meq/100 g
- Pretreatment of soil prior to use: drying at 80 °C for 24 h, sieving (2 mm mesh)

ADDITION OF INOCULUM TO SOIL SAMPLES: yes; prepared acc. to Lindhahl and Bakken (FEMS Microbiol. Ecol.,1995) (see above under 'Details on preparation and application of test substrate')

EFFECT PARAMETERS MEASURED after incubation period: soil nitrification, bacterial diversity (bacterial DNA), number of total protozoans (amoebae and heterotrophic flagellates) and heterotrophic flagellates

VEHICLE CONTROL PERFORMED: yes

RANGE-FINDING STUDY: no data

Nominal and measured concentrations:

Nominal concentrations: 1, 3, 10, 30, 100, 300, 1000, 3000 mg/kg soil dw;
Measured levels prior to test: 10 and 1000 mg/kg soil dw, recovery: 9.6 and 880 mg/kg soil dw;
Mean value for measured/nominal concentrations: 0.88 (mean recovery rate)

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

4 wk

Dose descriptor:

NOEC

Effect conc.:

26 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Effect concentrations 2

Key result

Duration:

4 wk

Dose descriptor:

EC10

Effect conc.:

42 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Remarks on result:

other: 95% C.I.: 0 - 70 mg/kg soil dw

Effect concentrations 3

Key result

Duration:

4 wk

Dose descriptor:

EC50

Effect conc.:

250 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Remarks on result:

other: 95% C.I.: 220 - 380 mg/kg soil dw

Effect concentrations 4

Duration:

4 wk

Dose descriptor:

other: EC5

Effect conc.:

2 400 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: total number of protozoa

Effect concentrations 5

Duration:

4 wk

Dose descriptor:

other: EC5

Effect conc.:

250 mg/kg soil dw

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: number of heterotrophic flagellates

Details on results:

Nitrogen mineralisation was generally the most sensitive of the three endpoints measured.
[Note: All substances affected nitrate production from > 20 - < 100 mg/kg soil dw, except acridine, which showed no substantial toxciity at ≥ 1000 mg/kg soil dw (Report Table 2)].

Reported statistics and error estimates:

Nitrate production: NOEC: ANOVA and Dunnett's procedure; EC10 and EC50: Linear interpolation
Protozoans: EC5 instead of NOEC: Three parameter relationship (by means of SigmaStat)