2-pyridylamine

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From December 17th, 2015 to January 14th, 2016.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source: A sample of activated sludge was taken from the aeration tank of Sewage Treatment Plant ”Czajka”, Warsaw, receiving predominantly domestic sewage.
- Preparation of inoculum: The coarse particles were removed by settling and the supernatant was discarded. The sludge was washed in the mineral medium. The concentrated sludge was suspended in mineral medium to yield a concentration of 3-5 g suspended solids/l and it was aerated, at the test temperature of 22 °C, until application next day. A sample was withdrawn just before use for the determination of the dry weight of the suspended solids.
- Amount of inoculum suspended: 30 mg/L.

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- pH: 7.12
- Test temperature: 22 ± 2°C
- Volume of test solution in flask, V: 0.164 L
- Initial test item concentration: 100 mg/L (51.3 mg/L of organic carbon)
- Composition of medium: 10ml of solution (A) in 800 ml water, plus 1 ml solutions (B), (C), (D) and 11ml water. The following stock solutions were used, prepared with analytical grade reagents:
- Solution (A) contains: 8.50 g monopotassium dihydrogen orthophosphate (KH2PO4), 21.75 g dipotassium monohydrogen orthophosphate (K2HPO4), 33.40 g disodium monohydrogen orthophosphate dihydrate (Na2HPO4·2H2O), 0.50 g ammonium chloride (NH4Cl), in 1 L double-distilled water.
- Solution (B) contains: 27.50 g calcium chloride, anhidrous (CaCl2) in 1L water.
- Solution (C) contains: 22.50 g magnesium sulphate heptahydrate (MgSO4·7H2O) in 1L water.
- Solution (D) contains: 0.25 g iron(III) chloride hexahydrate (FeCl3·6H2O) in 1L water.
- The water used is double-distilled, containing 3mg/L of organic carbon (< 10% of the organic carbon content introduced by the test item), checked by DOC analysis using spectrophotometer Hach DR 3900 and Hach-Lange reagents.
- Calculated ThOD(NH4): 1.70 mgO2/mg of test item; Calculated ThOD(NO3): 3.06 mgO2/mg test item.
- Suspended solids concentration: 30 mg/L

TEST SYSTEM
- Number of culture flasks/concentration: triplicates were used, flasks #13, 14, 15 containing test item (100 mg/l) and inoculum 30mg/L SS.
- Measuring equipment: O2 uptake was measured by a closed WTW OxiTop OC 110 repirometer. The data were read out every 112 min during the 28 day test (40 320 min that is 360 readings) and were recorded and stored in the measuring heads of the sample bottles.

CONTROL AND BLANK SYSTEM
- Inoculum blank: flasks #1,2, 3, containing only inoculum 30mg/L SS.
- Procedure control: flasks #10, 11, 12 containing reference item (sodium acetate 100 mg/l) and inoculum 30mg/L SS.
- Toxicity control: flasks #16, 17, 18 containing test item, reference item and inoculum 30mg/L SS.

STATISTICAL METHODS: The calculations and the graphs were performed using SigmaPlot 9.0 software of SYSTAT Software, Inc., USA purchased from GAMBIT CoiS Ltd, Poland.

Reference substance:

acetic acid, sodium salt

Remarks:

CAS No: 127-09-3, purity ≥ 98.5%, source: Eurochem.

Key result

Parameter:

% degradation (O2 consumption)

Value:

9.2

Sampling time:

28 d

Details on results:

As the test substance contains nitrogen, the observed oxygen uptake by the reaction mixture was corrected for the amount of oxygen used in oxidising ammmonium to nitrite and nitrate. After correction for oxygen uptake for interference by nitrification and by the blank inoculum control, the final aerobic biodegradation of the test item on the 28th test day was 9.2%.

Results with reference substance:

The reference item reached the pass level (60%) on day 6.

Table 1. Correction for oxygen uptake for interference by nitrification.

days	0			28			difference
1) Concentration of nitrate (mg N-NO3/l)	#13	#14	#15	#13	#14	#15	#13	#14	#15
	0.123	0.114	0.161	0.207	0.194	0.211	0.084	0.080	0.050
							0.071 ± 0.019
2) Oxygen equivalent (4.57× N-NO3) (mg/l)							0.324
3) concentration of nitrite  (mg N-NO2/l)	#13	#14	#15	#13	#14	#15	#13	#14	#15
	0.026	0.021	0.018	0.016	0.020	0.015	-0.010	-0.001	-0.003
							-0.005 ± 0.005
4) Oxygen equivalent (3.43×N-NO2) (mg/l)							-0.017
5) total oxygen equivalent 2) + 4)							0.31

 

Table 2. Sample oxygen uptake: biodegradability

		time, days
		3	5	7	9	12	14	16	18	21	23	25	28
Test item O2 uptake, mg/l	a1	3.2	6.2	9.5	13.0	17.9	22.0	24.7	28.4	34.0	38.6	41.4	45.2
	a2	6.1	10.9	15.6	19.1	25.9	30.3	34.0	36.9	43.6	47.5	50.7	54.9
	a3	4.6	8.2	14.0	18.8	25.8	29.6	32.1	34.5	39.6	43.1	47.2	49.5
	am. avg	4.6	8.4	13.0	17.0	23.2	27.3	30.3	33.2	39.1	43.1	46.5	49.9
Blank test O2 uptake. mg/l	b1	3.5	7.3	12.1	16.9	25.2	28.9	33.8	38.4	44.7	47.8	51.7	57.9
	b2	9.4	14.0	17.6	20.5	22.7	24.9	25.7	27.2	30.0	32.4	33.1	33.7
	b3	10.0	12.8	15.3	17.4	19.7	24.3	25.8	26.3	30.9	31.3	33.5	34.2
	avg	9.7	13.4	16.5	18.9	21.2	24.6	25.8	26.7	30.5	31.8	33.3	34.0
Reference item O2 uptake. mg/l	w1	44.9	57.5	67.3	73.8	81.6	86.6	90.7	93.1	95.7	98.5	101.1	103.7
	w2	47.2	59.3	68.0	73.9	80.5	84.9	86.4	87.2	90.0	90.3	92.8	96.4
	w3	44.4	55.8	63.2	71.0	76.3	81.0	84.9	88.0	92.4	92.8	95.4	95.7
	wm. avg	45.5	57.5	66.1	72.9	79.4	84.2	87.4	89.4	92.7	93.9	96.4	98.6
Toxicity control O2 uptake. mg/l	a4tox1	48.8	57.9	68.2	75.8	83.3	89.6	95.5	102.1	112.7	117.3	123.4	127.9
	a5tox2	46.9	56.8	64.6	70.9	79.6	86.3	92.3	95.7	104.2	107.3	110.0	114.3
	a6tox3	49.4	64.2	76.5	85.3	93.2	99.2	104.0	107.0	114.3	118.6	121.2	124.4
	toxm. avg	48.4	59.6	69.8	77.4	85.4	91.7	97.3	101.6	110.4	114.4	118.2	122.2
Corrected test item O2 uptake, mg/l	a1 - bm	-6.5	-7.3	-6.9	-5.9	-3.4	-2.5	-1.1	1.7	3.5	6.8	8.1	11.2
	a2 - bm	-3.6	-2.5	-0.9	0.2	4.7	5.8	8.2	10.1	13.2	15.7	17.4	20.9
	a3 - bm	-5.1	-5.2	-2.5	-0.1	4.5	5.1	6.4	7.7	9.2	11.2	13.9	15.6
Reference item % degradation ThOD = 0.78 mgO2/mg C = 100 mg/l	R 1(w1)	45.2	56.5	65.2	70.4	77.4	79.6	83.3	85.0	83.6	85.5	86.8	89.4
	R1 (w2)	48.1	58.8	66.0	70.6	75.9	77.3	77.8	77.5	76.3	75.0	76.3	80.0
	R3 (w3)	44.5	54.3	59.9	66.8	70.6	72.3	75.8	78.5	79.5	78.1	79.6	79.1
	Rtoxavg	45.9	56.5	63.7	69.2	74.6	76.4	78.9	80.4	79.8	79.5	80.9	82.9

*The result b1 is not taken into average value calculations, as it is distinctly greater than other blank test O2 uptakes.

 

Table 3. Toxicity test

days	0			28			difference
1) Concentration of nitrate (mg N-NO3/l)	#16	#17	#18	#16	#17	#18	#16	#17	#18
	0.171	0.152	0.131	0.259	0.241	0.212	0.088	0.089	0.081
							0.086 ± 0.004
2) Oxygen equivalent (4.57× N-NO3) (mg/l)							0.393
3) concentration of nitrite  (mg N-NO2/l)	#16	#17	#18	#16	#17	#18	#16	#17	#18
	0.020	0.031	0.018	0.014	0.018	0.012	-0.006	-0.013	-0.006
							-0.008 ± 0.004
4) Oxygen equivalent (3.43×N-NO2) (mg/l)							-0.027
5) total oxygen equivalent 2) + 4)							0.37

 

Table 4. The pH values of test flasks

flask #	13	14	15	1	2	3	10	11	12	16	17	18
	Test item			Control			Reference item			Toxicity test
initial	7.55	7.52	7.55	7.35	7.47	7.48	7.43	7.41	7.30	7.57	7.58	7.47
final	7.55	7.56	8.23	7.35	7.29	7.62	8.30	8.54	8.58	9.20	9.15	9.08

*No adjustment of pH was conducted

 

Validity criteria fulfilled:

yes

Remarks:

See remarks on results.

Interpretation of results:

not readily biodegradable

Conclusions:

The test item is not readily biodegradable. At the 28th day of the test the measured aerobic biodegradation of the test item attained 9.2%.

Executive summary:

The study of ready biodegradability of the test item in aerobic aqueous medium was performed with manometric respirometry method, according to OECD 301F / EU Method C.4 – D manometric respirometry  methods (GLP study). 30 mg/L of activated sludge were exposed to 100 mg/L of test item during 28 days, under aerobic conditions at 22ºC. Blank tests (only inoculum), and procedure tests with a reference item (sodium acetate) were run in parallel to check the operation of the procedures. To check the possible inhibitory effect of the test item, a toxicity test was run in parallel. The degradation was followed by the determination of oxygen uptake and measurements were taken at sufficiently frequent intervals to allow the identification of the beginning and end of biodegradation. As the test substance contains nitrogen, the observed oxygen uptake by the reaction mixture was corrected for the amount of oxygen used in oxidising ammmonium to nitrite and nitrate. After correction for oxygen uptake for interference by nitrification and by the blank inoculum control, the final biodegradation of the test item on the 28th test day was 9.2%. Therefore, the substance is not readily biodegradable under test conditions. In the toxicity test, a 35% biodegradation was attained after 28 days and, therefore, the test item is not assumed to be inhibitory.

Description of key information

Key study. Method according to OECD 301F / EU Method C.4 -D, GLP study. At the 28th day of the test the measured aerobic biodegradation of the test item attained 9.2%. Based on available information, the test item is not readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

A study of ready biodegradability of the testaccording to OECD 301F / EU Method C.4–D (GLP study) was performed. 30 mg/L of activated sludge were exposed to 100 mg/L of test item during 28 days, under aerobic conditions at 22ºC. Blank tests (only inoculum), and procedure tests with a reference item (sodium acetate) were run in parallel to check the operation of the procedures. To check the possible inhibitory effect of the test item, a toxicity test was run in parallel. After correction for oxygen uptake for interference by nitrification and by the blank inoculum control, the final biodegradation of the test item on the 28th test day was 9.2%. Therefore, the substance is not readily biodegradable under test conditions. In the toxicity test, a 35% biodegradation was attained after 28 days and, therefore, the test item is not assumed to be inhibitory.