Alkanes, C16-47, branched and linear

Toxicity to soil microorganisms

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil microorganisms

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

28 d

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/kg soil ww

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Remarks on result:

other: 95 % Cl not reported

Key result

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil ww

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Remarks on result:

other: 95 % CL limits not recorded

Details on results:

RESULTS
Range-finding Test
Absorbance values, nitrate concentrations and percentage inhibition of nitrogen transformation activity determined during the range-finding test are given in Table 1.
The results showed no significant effect on nitrogen transformation activity.
Based on this information a single test concentration of 1000 mg/kg soil was selected for the definitive test. This experimental design conforms to a “limit test” to confirm that at the maximum recommended concentration given in the Test Guidelines no effect on nitrogen transformation was observed.
Definitive Test
Absorbance values and nitrate concentrations determined on each sampling day are given in Table 2. The nitrate formation rates and percentage inhibition of nitrogen transformation activity on days 7 and 28 are given in Table 3.
The microbial biomass of the soil was shown to be 134 µg C/g (ISO 14240-2: 1997). This was equivalent to 1.4% of the total soil organic carbon content.
The variation between replicate control nitrification rates was less than 15% and therefore satisfied the validation criterion given in the Test Guidelines.
Accordingly the following results were determined from the data:
EC10 (28 days): >1000 mg/kg
EC25 (28 days): >1000 mg/kg
EC50 (28 days): >1000 mg/kg

where ECx is the test concentration that reduced soil nitrogen transformation activity by x %.
Statistical analysis of the nitrate concentration values was carried out for the control and test item group and showed no statistically significant decreases between the control and 1000 mg/kg test concentration (P≥0.05). Therefore the ‘No Observed Effect Concentration’ (NOEC) was 1000 mg/kg.
Temperature was maintained at approximately 21ºC throughout the test and the moisture content of each control and test vessel was checked on days 7, 14, 21 and 28 to ensure that the moisture content of the soil was maintained to 40% ± 5% of the WHC of the soil.
The mean nitrate formation rate in the range-finding test was significantly higher in the 1000 mg/kg test concentration than in the control; however inthe definitive test the mean nitrate formation rate in the control and 1000 mg/kg test concentration were similar.
The results in Tables 1 and 2 also show a significant increase in the nitrate formation rate between the control and 1000 mg/kg test vessels prepared in the range-finding and definitive test.  This increase may be due to the soil having been stored between the range-finding and definitive tests. The difference in nitrogen transformation rates between the range-finding and definitive tests is considered to have had no adverse effect on the study given that in both tests no inhibition of the nitrogen transformation rate between the control and test group was observed.

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

Statistical analysis
A Students t-test was carried out on the nitrate concentration values on Day 28 for the control and the 1000 mg/kg test concentration to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package (SAS 1999 - 2001).

Table 1: Absorbance Readings, Nitrate Concentrations and Inhibition of Nitrogen Transformation Activity from the Range-finding Test

Nominal Concentration (mg/kg soil)		Day 0		Nominal Concentration (mg/kg soil)		Day 28
		Absorbance* (324 nm)	Nitrate Concentration (mg NO3/kg)			Absorbance* (324 nm)	Nitrate Concentration (mg NO3/kg)	Mean Nitrate Formation Rate (mgNO3/kg/day)	% Inhibition
Control	R1	0.245	88.35	Control	R3	0.359	128.50	1.33	1
	R2	0.244	88.00		R4	0.342	122.50
10	R1	0.242	87.30	10	R3	0.359	128.50	1.52	[14]
	R2	0.235	84.85		R4	0.360	128.85
100	R1	0.251	90.50	100	R3	0.367	131.30	1.59	[20]
	R2	0.248	89.40		R4	0.385	137.65
1000	R1	0.245	88.35	1000	R3	0.615	218.55	5.01	[277]
	R2	0.248	89.40		R4	0.675	239.70

 

*Corrected for blank absorbance value

[Increase in nitrate formation rate as compared to controls]

 R₁-R₄= Replicates R₁to R₄

Table 2: Absorbance and Nitrate Concentrations from the Definitive Test

Nominal Concentration (mg/kg soil)		Day 0
		Absorbance* (324 nm)	Nitrate Concentration (mg NO3/kg)
Control	R1	0.304	109.15
	R2	0.295	105.95
	R3	0.291	104.55
	Mean	-	106.55
	CV%	-	2
1000	R1	0.313	112.30
	R2	0.306	109.85
	R3	0.310	111.25
	R4	0.295	105.95
	R5	0.306	109.85
	R6	0.307	110.20
	Mean	-	109.90
	CV%	-	2

 

Nominal Concentration (mg/kg soil)		Day 7
		Absorbance* (324 nm)	Nitrate Concentration (mg NO3/kg)
Control	R4	0.543	193.25
	R5	0.530	188.65
	R6	0.510	181.60
	Mean	-	187.33
	CV%	-	3
1000	R7	0.524	186.55
	R8	0.554	197.10
	R9	0.505	179.85
	R10	0.519	184.80
	R11	0.534	190.05
	R12	0.515	183.40
	Mean	-	186.96
	CV%	-	18

*Corrected for blank absorbance value

R₁-R₁₂= Replicates 1 to 12

CV = Coefficient of Variation

Table 2 (continued): Absorbance Readings and Nitrate Concentrations from the Definitive Test

Nominal Concentration (mg/kg soil)		Day 28
		Absorbance* (324 nm)	Nitrate Concentration (mg NO3/kg)
Control	R7	1.194	422.30
	R8	1.188	420.20
	R9	1.192	421.60
	Mean	-	421.37
	CV%	-	<1
1000	R13	1.192	421.60
	R14	1.198	423.75
	R15	1.193	421.95
	R16	1.175	415.65
	R17	1.232	435.70
	R18	1.199	424.10
	Mean	-	423.79
	CV%	-	2

*Corrected for blank absorbance value

 R₇-R₉= Replicates 7 to 9

 R₁₃-R₁₈= Replicates 13 to 18

CV = Coefficient of Variation

Table 3: Inhibition of Nitrogen Transformation Activity

Nominal Concentration (mg/kg)	Day 7		Day 28
	Nitrate Formation Rate (mg NO3/kg/day)	% Inhibition	Nitrate Formation Rate (mg NO3/kg/day)	% Inhibition
Control	11.61	-	11.24	-
1000	11.01	5	11.21	0

Validity criteria fulfilled:

yes

Conclusions:

The effect of the read across substance GTL Base Oil Distillates on the nitrogen transformation activity of soil microorganisms has been investigated over a 28-Day period and gave an EC50 value of greater than 1000 mg/kg. Correspondingly the No Observed Effect Concentration was 1000 mg/kg.

Executive summary:

Introduction.

A study was performed to assess the long-term effect of the read across substance ‘Distillates (Fischer-Tropsch), heavy, C18-50 - branched, cyclic and linear' after a single exposure, on the nitrogen transformation activity of soil microorganisms. The method followed that described in the OECD Guidelines for Testing of Chemicals (2000) No 216 “Soil Microorganisms: Nitrogen Transformation Test” referenced as Method C.21 of Commission Regulation (EC) No. 440/2008; EPPO (1994) Decision Making Scheme for the Environmental Risk Assessment of Plant Protection Products, Chapter 7: Soil Microflora, EPPO Bulletin 24 and SETAC-Europe (1995) Procedures for Assessing the Environmental Fate and Ecotoxicity of Pesticides.

Methods.

Following a preliminary range-finding test soil microorganisms were exposed to the test item at a single concentration of 1000 mg/kg for 28 days at a temperature of approximately 21 ºC, in the dark with the addition of powdered Lucerne-green-grass meal to act as a respiratory substrate.

The inhibitory effect of the test item on nitrogen transformation was assessed by the determination of nitrate concentration in the soil samples on Days 0, 7 and 28 and compared to data obtained from control soil samples.

Results.

The effect of the test item on the nitrogen transformation activity of soil microorganisms has been investigated over a 28-Day period and gave an EC₅₀ value of greater than 1000 mg/kg. Correspondingly the No Observed Effect Concentration was 1000 mg/kg.

CONCLUSION

The effect of the test item on the nitrogen transformation activity of soil microorganisms has been investigated over a 28-Day period and gave an EC₅₀ value of greater than 1000 mg/kg. Correspondingly the No Observed Effect Concentration was 1000 mg/kg.

Description of key information

- EC50 (28d): >1000 mg/kg dw test mat. (nominal) based on: nitrate formation rate
- NOEC (28d): ≥1000 mg/kg dw test mat. (nominal) based on: nitrate formation rate

Additional information

A nitrogen transformation test is available for the read across substance GTL Base Oil Distillates. The test was conducted in accordance with OECD 216 and GLP.

Soil microorganisms were exposed to the substance at concentrations of 1000 mg/kg (dry weight) for 28 days at a temperature of 21°C, in the dark, on natural soil with an organic carbon content of 0.97 ± 0.07% with addition of Lucerne-green-grass meal as a respiratory substrate. The inhibitory effect of the test substance on nitrogen transformation was determined by measurement of nitrate concentrations in soil samples taken on Days 7, 14, 21 and 28, in comparison to control samples.

The effect of the test substance on nitrogen transformation gave an EC₅₀greater than 1000 mg/kg dw.