Extract of fava d'anta, obtained from the fruits of Dimorphandra mollis (Leguminosae) by solvent extraction

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

toxicity to microorganisms, other

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16.03.2021-20.04.2021

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Test method according to OECD 301F. GLP study.

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 100 mg/L
- Sampling method: DOC analysis using spectrophotometer Hach DR 3900 and Hach-Lange reagents.
- Sample storage conditions before analysis: samples were analysed immediately after sampling.

Vehicle:

no

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: A sample of activated sludge was taken from the aeration tank of Sewage Treatment Plant ”Czajka” , Warsaw, receiving predominantly domestic sewage
- Name and location of sewage treatment plant where inoculum was collected: ”Czajka” , Warsaw, Poland
- Preparation of inoculum for exposure: Pre-conditioning consisted of 7 days aeration of activated sludge at the test temperature of 22 ± 1ºC.
- Pretreatment: The coarse particles were removed
- Initial biomass concentration: 30 mg/L SS

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

28 d

Test temperature:

22±1ºC

pH:

7.4±0.2

Details on test conditions:

TEST SYSTEM
- Test vessel: respirometer flasks.
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: glass, volume of test solution in flask, V: 0.164 L
- No. of vessels per concentration (replicates): 3, each containing 100 mg/L test item and 30 mg/L SS inoculum.
- No. of vessels per control (replicates):
Inoculum blank: 3 replicates, each containing 30 mg/L SS inoculum.
Procedure control: 3 replicates, each containing 100 mg/L reference item (sodium acetate) and 30 mg/L SS inoculum.
Toxicity control: 2 replicates, each containing 100 mg/L test item, 100 mg/L reference item (sodium acetate) and 30 mg/L SS inoculum.
- Sludge concentration (weight of dry solids per volume): 30 mg/L.
- Nitrification inhibitor used (delete if not applicable): none.

TEST MEDIUM / WATER PARAMETERS
- Preparation of test solutions: The solutions of the test and reference items were prepared, in separate batches, in mineral medium equivalent to a concentration of 100 mg test item/litre. The requisite volume of solutions of test and reference items, were introduced respectively, into triplicate flasks. 16.4 mg of test item and 16.4 ml of stock solution of the reference item were introduced into each of the triplicate right flasks. Mineral medium only was added to further flasks (for inoculum controls). Potassium hydroxide solution was added to each of the CO2-absorber compartments. The determined content of inoculum at the end of preconditioning was equal to 5.10 mg suspended solids/l. To give a concentration of suspended solids equal to 30 mg/l in each flask, 1 ml of preconditioned suspension of inoculum was added into each flask. The flasks were made up to 164 ml volume with prepared mineral medium.

OTHER TEST CONDITIONS
- Adjustment of pH: no.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Biodegradation % based on oxygen consumption.

TEST CONCENTRATIONS
- Test concentrations: 100 mg/L

Reference substance (positive control):

yes

Remarks:

Acetic acid, sodium salt. CHEMPUR, purity p.a. 99.7%

Key result

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Readily biodegradability

Details on results:

The NOEC of the test item was determined to be 100 mg/L since the substance degrades well (87.5% of biodegradation at 28 days) and did not inhibit the biodegradation in the toxicity test (69.4% of biodegradation after 14 days) up to 100 mg/L.

Results with reference substance (positive control):

- Results with reference substance valid: yes
- The reference item reached 74.8 % of biodegradation and the level for ready biodegradability by 5 days (pass level: biodegradation higher than 60% before day 14)

Table 2. Sample oxygen uptake: biodegradability.

		time, days
		1	3	5	7	9	12	14	16	18	21	23	25	28
Test item O2 uptake, mg/l	a1	6.1	40.1	51.8	60.9	67.9	81.3	87.5	91.5	94.4	102.2	104.7	107.4	113.7
	a2	5.5	39.3	51.8	64.0	74.9	87.7	93.7	99.1	101.8	107.4	109.0	110.4	114.4
	a3	6.3	41.9	53.9	62.7	70.5	80.2	82.5	84.7	85.7	87.5	87.2	86.2	86.3
	am. avg	5.8	39.7	51.8	62.4	71.4	84.5	90.6	95.3	98.1	104.8	106.9	108.9	114.0
Blank test O2 uptake, mg/l	b1	3.0	9.4	11.5	13.2	15.5	17.0	17.8	19.1	21.0	22.8	24.5	26.3	28.0
	b2	5.9	10.5	11.4	14.7	15.3	16.7	18.2	20.8	20.3	21.8	22.4	24.4	23.9
	b3	4.3	9.2	8.5	13.1	15.2	15.6	17.2	18.4	20.4	22.3	24.5	27.3	26.8
	bm avg	4.4	9.7	10.5	13.7	15.3	16.4	17.7	19.4	20.6	22.3	23.8	26.0	26.3
Reference item O2 uptake, mg/l	w1	34.7	48.0	53.6	60.6	63.5	68.1	70.4	71.4	72.7	75.8	78.0	80.0	80.3
	w2	17.1	53.2	67.4	75.2	79.4	86.2	87.2	90.4	93.1	95.8	96.2	99.1	101.3
	w3	16.9	50.1	61.9	67.8	73.4	78.0	80.2	81.7	82.7	84.9	86.8	87.3	88.9
	wm.avg	25.8	49.1	57.8	64.2	68.5	73.1	75.3	76.5	77.7	80.3	82.4	83.6	84.6
Toxicity control O2 uptake, mg/l	tox1	26.1	61.8	84.6	115.2	129.4	141.2	148.7	162.4	169.8	180.5	186.5	190.7	196.7
	tox2	21.4	55.0	73.9	93.1	107.8	124.6	134.3	142.8	152.4	160.4	166.5	168.6	174.4
	tox.avg	23.8	58.4	79.3	104.1	118.6	132.9	141.5	152.6	161.1	170.4	176.5	179.6	185.6
Corrected test item O2 uptake, mg/l	a1 - bm	1.7	30.4	41.4	47.2	52.6	64.9	69.8	72.1	73.8	79.8	80.9	81.4	87.4
	a2 - bm	1.1	29.6	41.4	50.3	59.5	71.3	76.0	79.7	81.2	85.1	85.2	84.4	88.1
	a3 - bm	-	-	-	-	-	-	-	-	-	-	-	-	-
Reference item % degradation ThOD = 0.78 mgO2/mg C = 100 mg/l	R1 (w1)	38.8	49.2	55.3	60.2	61.8	66.2	67.5	66.6	66.8	68.6	69.4	69.2	69.3
	R2 (w2)	-	-	-	-	-	-	-	-	-	-	-	-	-
	R3 (w3)	16.0	51.9	66.0	69.4	74.4	79.0	80.0	79.8	79.6	80.2	80.8	78.6	80.3
	Ravg	27.4	50.5	60.7	64.8	68.1	72.6	73.8	73.2	73.2	74.4	75.1	73.9	74.8±7.8
Test item % degradation COD = 141 mgO2/l	R1 (a1)	1.7	30.4	41.2	47.1	52.4	64.7	69.6	71.8	73.6	79.6	80.7	81.2	87.2
	R2 (a2)	1.1	29.5	41.2	50.2	59.4	71.1	75.8	79.4	80.9	84.8	84.9	84.2	87.8
	R3 (a3)	-	-	-	-	-	-	-	-	-	-	-	-	-
	Raavg	1.4	29.9	41.2	48.6	55.9	67.9	72.7	75.6	77.3	82.2	82.8	82.7	87.5±0.4
Toxicity test % degradation	R1 (tox1)	12.2	29.2	41.6	56.9	64.0	70.0	73.5	80.2	83.7	88.7	91.3	92.4	95.6
	R2 (tox2)	9.5	25.4	35.6	44.6	51.9	60.7	65.4	69.2	73.9	77.5	80.0	80.0	83.1
	Rtoxavg	10.9	27.3	38.6	50.7	57.9	65.3	69.4	74.7	78.8	83.1	85.6	86.2	89.4±8.8

 

Table 3. The pH values of test flasks

flask #	1	2	3	4	5	6	7	8	9	10	11
	Inoculum blank			Reference item			Test item			Toxicity test
initial	7.37	7.41	7.44	7.44	7.46	7.46	7.45	7.45	7.46	7.44	7.43
final	7.43	7.42	7.32	9.26	8.93	8.91	7.68	7.59	7.51	8.38	8.45

 

Validity criteria fulfilled:

yes

Remarks:

See "overall remarks".

Conclusions:

The NOEC (28d) for microorganism toxicity of the test item was determined to be 100 mg/L (based on biodegradation).

Executive summary:

A ready biodegradability manometric respirometry test was perfomed according to OECD Guideline 301 F, GLP study. The test item was incubated for 28 days at a concentration of 100 mg/L with 30 mg/L SS activated sludge inoculum under aerobic conditions in a mineral medium. Additionally, to check the possible inhibitory effect of the test item, a toxicity test was run in parallel (100 mg/L test item with 100 mg/L reference substance and 30 mg/L SS activated sludge inoculum) at the same conditions. The NOEC (28d) of the test item was determined to be 100 mg/L since the substance degraded well (87.5% of biodegradation at 28 days) and did not inhibit the biodegradation in the toxicity test (69.4% of biodegradation after 14 days) up to 100 mg/L.

Description of key information

Key study: Test method according to OECD 301F, GLP study. The NOEC (28d) for microorganism toxicity of the test item was determined to be 100 mg/L.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

100 mg/L