2-benzyl-2-dimethylamino-4'-morpholinobutyrophenone

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-05-04, 2009-06-12

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

yes

Remarks:

The air temperature in the climate chamber increased and decreased temporarily for short time periods up to 23 °C and down to 17 °C (recommended 20 ± 2 °C. 500 g artificial soil were used instead of 750 g.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

- Method of mixing into soil: The test item was mixed in an amount of quartz sand sufficient to prepare a mixture for each test item concentration separately. An appropriate amount of these test item/quartz sand mixtures served to prepare the different concentrations of the test item in the artificial soil. The test item was incorporated into the remainder of the artificial soil by thoroughly mixing in the respective test item/quartz sand mixtures for approx. 10 minutes as described in the test guideline.
- Controls: Untreated deionised water control

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

TEST ORGANISM
- Common name: Earthworm
- Source: ECT Ökotoxikologie GmbH
- Age at test initiation: adult worms (with clitellum)
- Weight at test initiation: 300 - 600 mg

ACCLIMATION
- Acclimation period: 24 h
- Acclimation conditions: same as test

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

14 d

Post exposure observation period:

None

Test temperature:

17 - 23 °C

pH:

6.0 - 6.2

Moisture:

54.9 - 57.1 % of WHCmax

Details on test conditions:

TEST SYSTEM
- Test container (material, size): 1.5 L preservation jars ("Tulpenform"), Co. Weck, Wehr-Öflingen. The test vessels were covered by glass lids.
- Amount of soil or substrate: 500 g (dw)
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 4

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Composition (if artificial substrate): 5% Sphagnum peat (no visible plant remains; finely ground and air-dried); 20% Kaolin-Clay (kaolinite content > 30 %); 68 - 69 % Quartz sand (fine sand content with particles between 50 and 200 µm higher than 50 %); ca. 1 % Calcium carbonate (CaCO3)
- Maximum water holding capacity (in % dry weigth): 54.2
- Pretreatment of soil: One day before starting the test the artificial soil (per replicate 500 g dw) was premoistened with deionised water (20 - 40 % of WHCmax)

OTHER TEST CONDITIONS
- Photoperiod: Permanent
- Light intensity: 449 - 558 Lux

EFFECT PARAMETERS MEASURED: Mortality and biomass determined after 7 and 14 days

TEST CONCENTRATIONS: Limit test with two concentrations
- Spacing factor for test concentrations: 10
- Justification for using less concentrations than requested by guideline: Previous Range-Finding Test

Nominal and measured concentrations:

Nominal: 100 and 1000 mg test item/kg soil dw (dry weight)

Reference substance (positive control):

yes

Remarks:

10, 20, 40, and 80 mg chloroacetamide/kg

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Biomass

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Biomass

Details on results:

- Mortality at end of exposure period: None, table 1
- Total mass of adults at beginning of test: Table 2
- Changes in body weigth of live adults (% of initial weight) at end of exposure period: Table 1
- Morphological abnormalities: None
- Behavioural abnormalities: None
- Other biological observations: None

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: LC50=50.2 mg/kg

Reported statistics and error estimates:

R/s test procedure confirmed the correspondence with normal distribution. The hypothesis of homogeneity of the biomass data was accepted by the Cochran's test. Therefore, the pairwise comparison of the concentrations with the control test by the Student-t test was used for the statistical analysis.
The ANOVA showed no significant difference but the Student-t test (2-sided, p ≤ 0.05) showed a significant difference in biomass development of individual adults after 14 days between the control and the highest concentrations of the test item tested (i.e. 1000 mg test item/kg soil (dw)) (table 3).
Therefore, the NOEC(Biomass) was determined as 100 mg test item/kg soil (dw) and accordingly, the LOEC(Biomaass) as 1000 mg test item/kg soil (dw).

Table 1. Mean live weight per surviving worm at all concentrations and the control at the beginning of the test, after 7 and 14 days (n = 40). Data are given as absolute values (biomass) and in percent of the initial weight

Concentration [mg test item/kg soil (dw)]	Mortality [%]	Biomass [% of initial weight]
Control	0.0	110.3
100	0.0	104.5
1000	0.0	103.1*

* significantly different to control (Student-t test; p ≤ 0.05, 2-sided)

Table 2. Number and biomass (mg) of the earthworms determined at the beginning, after 7 and 14 days of the test. The concentration is given in mg test item/kg soil (dw) (SD = Standard Deviation; I = Individual Weight).

Day	Parameter	Control				100				1000
		A	B	C	D	A	B	C	D	A	B	C	D
0	Weight ( l )	516	306	321	303	460	307	335	386	302	302	336	360
		392	371	318	396	444	388	469	325	384	319	401	317
		347	336	422	363	436	310	328	328	418	383	397	442
		348	303	456	403	338	314	347	377	337	552	407	451
		411	307	344	386	391	345	497	410	370	337	320	301
		378	328	465	485	387	480	438	369	529	313	485	351
		315	397	359	305	381	356	300	413	452	325	358	384
		412	362	455	302	321	343	352	346	302	324	359	355
		308	325	316	434	342	306	371	370	317	303	334	364
		421	310	321	324	316	311	333	312	415	301	375	367
	Number	10	10	10	10	10	10	10	10	10	10	10	10
	Average	384.8	334.5	377.7	370.1	381.6	346	377	363.6	382.6	345.9	377.2	369.2
	SD	61.1	32.1	64.1	62.2	52.2	54.3	66.8	35.1	73.1	76.3	48.3	47.4
7	Weight (Sum)	4040	3800	4222	4156	3931	3870	3926	3861	3999	3738	3985	3549
	Number	10	10	10	10	10	10	10	10	10	10	10	10
	Average	404	380	422.2	415.6	393.1	387	392.6	386.1	399.9	373.8	398.5	354.9
14		A	B	C	D	A	B	C	D	A	B	C	D
	Weight ( l )	419	388	350	408	387	360	476	448	388	362	502	333
		460	445	494	350	475	386	389	444	515	351	377	333
		466	462	535	412	399	355	318	381	389	570	418	321
		372	403	393	470	355	518	474	400	393	400	426	296
		396	378	370	509	387	337	372	468	358	361	363	394
		376	386	494	378	316	318	333	388	436	350	380	321
		341	340	422	499	472	383	478	363	390	331	382	463
		389	370	369	395	325	352	370	345	251	307	396	441
		501	336	356	303	291	419	358	354	362	344	370	379
		389	355	372	33 1	360	321	344	326	315	402	365	371
	Number	10	10	10	10	10	10	10	10	10	10	10	10
	Average	410.9	386.3	415.5	405.5	376.7	374.9	391.7	391.7	379.7	377.8	397.9	365.2
	SD	49.9	41.4	67.6	69.7	61.4	59	47.9	47.9	69.5	73.3	42.3	54.9

Table 3. STUDENT t-test for Homogeneous Variances.

Pair-wise comparison of treatments with "Control" by the t-test procedure. Significance was Alpha = 0.05, two-sided; Mean: arithmetic mean; n: sample size; s²: variance; %MDD: minimum detectable difference to Control (in percent of Control); t: sample t; p(t): probability of sample t for Ho: µ1 = µ2; the differences are significant in case p(i) ≤ Alpha ; p(F): probability of F computed by a F-test (Ho: varl = var2 (homogeneity); p(F) > 0.05 is the criterion of variance homogeneity. ((Control(c) and treatment(t) variance was applied; df = N - k; N: sum of treatment replicates n(i); k: number of treatments).

Treatm. [mg/kg]	Mean	s^2	df	%MDD	t	p(t)	Sign.	p(F)
Control	37.775	112.882
100.00	16.700	249.787	6	61.7	-2.21	0.069	-	0.266
1000.00	11.425	316.129	6	67.1	-2.54	0.044	+	0.210

+ = significant; - = non-significant

Validity criteria fulfilled:

yes

Conclusions:

No toxic effects have been observed in the study with soil living organisms. Statistically significant bodyweight changes were noted in the 1000 mg/kg dw/soil group.

Executive summary:

In this guideline (OECD 207) study conducted with GLP certification, the test material (EC 404-360-3) was determined to not be toxic to soil macroorganisms (worms). The test material (100 mg/kg dw/soil and 1000 mg/kg dw/soil) was exposed to annelid worms (Eisenia fetida) for 14 days. No treatment related mortality was observed during the test. Statistically significant bodyweight changes were noted in the 1000 mg/kg dw/soil group. The NOEC was set at 100 mg/kg dw/soil, as the lowest treated group without effects.

Description of key information

Study conducted to recognised training guidelines with GLP

Additional information

The acute toxicity of the test item to the earthworm Eisenia fetida was tested in a GLP guideline study according OECD 207 (ECT Oekotoxikologie GmbH 2009). After 14 d no mortality was observed at the control and at the tested limit concentration of 100 and 1000 mg test item/kg artificial soil dry weight. The biomass of individual adults after 14 days amounted to 110.3 % of the initial weight in the control, 104.5 % at 100 mg test item/kg soil (dw) and 103.1 % at 1000 mg test item/kg soil (dw). Therefore, the NOEC_biomass was determined as 100 mg test item/kg soil (dw) and accordingly, the LOEC_Biomass as 1000 mg test itemlkg soil (dw).