LBX98

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: inherent biodegradability

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

weight of evidence

Justification for type of information:

A 301D study with the target substance showed biodegradation of 39 % in 28 days. However, this test is very stringent and it does not necessarily mean that the substance would not biodegrade under environmental conditions. The 2,3 xylenol and 2,4 xylenol were shown to be inherently degradable in a test similar to OECD guideline 302B with a removal of 89.3 -97.5 % after 5 days of incubation . The 2,3 -xylenol and 2,4 -xylenol degradation rates were >15 mg COD/g/h which fulfilled the criteria for being readily biodegradable. Ethyl phenols are considered to be readily biodegradable with 72% degradation after 7 days, and m-, p- and ocresol were shown to be inherently degradable in a test similar to OECD guideline 302 B with degradation rates >15 mg COD/g/h and removal of 95 -96 % of m-cresol after 5 days of incubation. Based on all available information for cresols, ethyl-phenols and xylenols the target substance is described as inherently biodegradable.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 302 B (Inherent biodegradability: Zahn-Wellens/EMPA Test)

Deviations:

yes

Remarks:

minor deviations

Principles of method if other than guideline:

On the basis of experience obtained the Department of Water Technology and Environmental Engineering has developed a standard test for the comparison of biological degradability of organic substances.

GLP compliance:

no

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):Biological medium prepared by adding 1 mL each of solutions of calcium chloride (25g in 1L distilled water), magnesium sulphate (22.5 g in 1L) and ferric chloride (0.25g in 1L) with 5 ml ammonium sulphate solution (10g in 1 L), 20 ml of pH 7.2 phosphate buffer and 100 ml tapwater to ca 800 ml of distilled water, then made up to 1000 ml with distilled water.

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, adapted

Details on inoculum:

100 mg/L dry weightActivated sludge taken from sewage treatment plant is cultivated in a 1000ml volumetric cyclinder and aerated with pressure air. Every day 200 ml of the mixture is driven off so that the sludge age is 5 days. After driving off the 200 ml aeration is interrupted, and after sedimentation ca 600 ml of the liquid phase is driven off. The residue (200 ml of thickened activated sludge) is diluted to ca 800 ml and 600 mg/l starch or glucose, 600 mg/L peptone, 25 mL phosphate buffer pH 7.2 and the solution of the tested compound are added. Then the mixture is made up to 1000 ml with tap water and aerated for 23 h. After this period the procedure is repeated. The concentration of the tested substance is gradually increased so that after 20 days of adaptation it reaches the equivalent value of 200 mg/L COD.

Duration of test (contact time):

ca. 1 - ca. 20 d

Initial conc.:

200 mg/L

Based on:

COD

Parameter followed for biodegradation estimation:

other: COD removal

Details on study design:

Test compound was sole source of carbon. To 1000-1500ml of the biological medium such amount of the solution of the substance tested is added that the intial COD is 200 mg/L. Then an amount of the adapted activated sludge is dosed to the medium that the concentration of the dry matter is 100 mg/L. Simultaneouly a blank test is prepared to verify the activity of the inoculum. With volatile substances a test without the inoculum is carried out as well to differentiate the actual biological degradation and the losses due to volatilization. 50-80 ml of the mixture are taken, and after filtration through a medium porosity filter paper, the inital vlaues of COD or organic carbon of the liquid phase are determined. Initial levels in the beakers are marked and afterwards placed in a dark room at 20 +/- 3 degrees C on magnetic stirrers. The oxygenation capacity of this device was ca 11 mg oxygen h/L at 800 rev/min. At regular time intervals ca 50-80 ml of the sample are taken for analysis. The experiment was carried out until there is no decrease in COD. After that time the total percentage of COD removed and the rate of degradation are evaluated. If 90% of the initial COD is removed in 120h of incubation (degradation rate being more than 15 mg of COD g/h) then the test substance can be considered to be readily biodegradable.

Parameter:

other: COD removal

Value:

95.5

Sampling time:

5 d

Remarks on result:

other: m-cresol

Parameter:

other: COD removal

Value:

95

Sampling time:

5 d

Remarks on result:

other: o-cresol

Parameter:

other: COD removal

Value:

96

Sampling time:

5 d

Remarks on result:

other: p-cresol

Parameter:

other: COD removal

Value:

89.3

Sampling time:

5 d

Remarks on result:

other: 3,5-xylenol

Parameter:

other: COD removal

Value:

95.5

Sampling time:

5 d

Remarks on result:

other: 2,3-xylenol

Details on results:

Initial degradation rate: (95.5%) 55.0 mg COD/g/h (m-cresol); (95%) 54.0 mg COD/g/h (o-cresol); (96%) 55.0 mg COD/g/h (p-cresol); (89.3%) 11.0 mg COD/g/h (3,5-xylenol); (95.5%) 35.0 mg COD/g/h (2,3-xylenol); (94.5%) 28.2 mg COD/g/h (2,4-xylenol); (97.5%) 13.4 mg COD/g/h (3,4-xylenol); (94.5%) 10.6 mg COD g/h (2,5-xylenol); (94.3%) 9.0 mg COD/g/h (2,6-xylenol).

Substance name	Amount removed (%)	Degradation rate mg/COD/g/h
m-cresol	95.0	55.0
p-cresol	96.0	55.0
o-cresol	95.0	54.0
3,5 -xylenol	89.3	11.1
2,3 -xylenol	95.5	35.0
2,4 -xylenol	94.5	28.2
3,4 -xylenol	97.5	13.4
2,5 -xylenol	94.5	10.6
2,6 -xylenol	94.3	9.0

Interpretation of results:

inherently biodegradable

Executive summary:

The author carried out experiments on the degree and rate of biological degradation of 123 organic compounds with respect to the decrease of organic substance in terms of COD. The organic substances were the sole source of carbon for the microbes of the inoculum, adapted activated sludge being the inoculum. Samples were taken at regular intervals and the COD measured until there was no more decrease in COD. After that time the total percentage of COD removed and the rate of degradation were evaluated.

m-, p- and o-Cresol is inherently degradable as shown by a test similar to OECD guideline 302 B with a removal of 95 -96 % of m-cresol after 5 days of incubation. The degradation rates were >15 mg COD/g/h which was the criteria for being readily biodegradable.

The xylenols are inherently degradable as shown by a test similar to OECD guideline 302 B with a removal of 89.3 -97.5 % after 5 days of incubation . 2,3 -xylenol and 2,4 -xylenol had degradation rates >15 mg COD/g/h which was the criteria for being readily biodegradable (Pitter 1976).

The above results are suitable to be used for read across for Reaction mass of 2,4-xylenol and 2,5-xylenol, due to the category being based on structural similarity and comparable physicochemical properties, leading to similar (eco)toxicological properties.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

weight of evidence

Rationale for reliability incl. deficiencies:

other:

Justification for type of information:

A 301D study with the target substance showed biodegradation of 39 % in 28 days. However, this test is very stringent and it does not necessarily mean that the substance would not biodegrade under environmental conditions. The 2,3 xylenol and 2,4 xylenol were shown to be inherently degradable in a test similar to OECD guideline 302B with a removal of 89.3 -97.5 % after 5 days of incubation . The 2,3 -xylenol and 2,4 -xylenol degradation rates were >15 mg COD/g/h which fulfilled the criteria for being readily biodegradable. Ethyl phenols are considered to be readily biodegradable with 72% degradation after 7 days, and m-, p- and ocresol were shown to be inherently degradable in a test similar to OECD guideline 302 B with degradation rates >15 mg COD/g/h and removal of 95 -96 % of m-cresol after 5 days of incubation. Based on all available information for cresols, ethyl-phenols and xylenols the target substance is described as inherently biodegradable.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I))

Deviations:

yes

Remarks:

Voith sapromat B12

GLP compliance:

no

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

Minicipal sewage sludge with initial cell concentration: 30 mg/l

Duration of test (contact time):

>= 20 - <= 40 d

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Reference substance:

aniline

Remarks:

100 mg/L

Parameter:

% degradation (O2 consumption)

Value:

>= 80 - <= 95

Sampling time:

40 d

Remarks on result:

other: readily biodegradable

Details on results:

The oxygen uptake curves are not reported. However, the authors state that all test compounds revealed the lag phase, biodegradation phase and the plateau region within a period of 10 days, indicating that the 10-day window criteria is met.first order biodegradation constant (hr-1): ln k = -5.87maximum specific substrate uptake rate per unit biomass km = 18.5 / day (Aniline 16.1, Phenol 16.9)p-Cresol is slightly better biodegradable than phenol and aniline

Results with reference substance:

degradation of reference substance aniline >=60 % within 28 days

Incubation period 20-40 days; no oxygen uptake curve given

Interpretation of results:

readily biodegradable

Executive summary:

In a test according to OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I)), 80 to 95 % of p-cresol were degraded within 40 days. p-Cresol was slightly better biodegradable than phenol and aniline supporting the conclusion that it is readily biodegradable (Desai, Govind, and Tabak, 1990).

The above results are suitable to be used for read across for Reaction mass of 2,4-xylenol and 2,5-xylenol, due to the category being based on structural similarity and comparable physicochemical properties, leading to similar (eco)toxicological properties.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Deviations:

no

GLP compliance:

yes

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

Secondary effluent from a predominantly domestic STP. Collected 15 June 2010 and filtered. Concentration of 1ml/L.

Duration of test (contact time):

>= 28 d

Parameter followed for biodegradation estimation:

DOC removal

Remarks:

BOD determined

Details on study design:

Replicate standard BOD bottles were filled with the test concentration, prepared by addition of a sample stock solution to inoculated mineral medium. To check the intergrity of the test procedure a readily degradable reference chemical, sodium acetate, was tested and blank tests set up. Initial DO concentrations were measured and then incubated in the dark at ca 20 degrees C. Further measuresments of DO were made on bottles removed after 7, 14, 21 and 28 days. The theoretical OD was calculated based on information provided by the client and structures sourced from the Internet.

Reference substance:

other: Sodium acetate

Test performance:

The reference material was readily degradable at 71% degradation in < 10 days.

Key result

Parameter:

other: calculated BOD

Value:

39

Sampling time:

28 d

Details on results:

The xylenol mixture failed to meet the requirements for a pass in this test (>60% degradation relative to the COD value - a maximum of 39% was recorded). The ThOD, calculated based on information provided by the client and structures sourced from the internet, has a value of 2623 mg O2/g for HPV Xylenols.

Results with reference substance:

The reference material was readily degradable at 71% degradation in < 10 days.
The blank dissolved oxygen loss was 0.42mg/l (Guideline requires blank dissolved oxygen los of <1.5 mg/l).

	Incubation period (days)
	0	7		14		21			28
Incubator temperature (°C)	20.5	20.5		20.5		20.6			20.5
Average blank - DO (mg/l)	8.57	8.51		8.44		8.36			8.15
Replicate number	1	1	2	1	2	1	2	3	1	2	3
Test substance – DO (mg/l)	8.57	6.91	7.00	6.27	6.87	6.03	6.05	5.82	5.79	5.86	5.42
Test substance - BOD (mg O2/g)	-	658	621	893	646	959	951	1045	971	942	1123
Reference material – DO (mg/l)	8.57	5.65	5.82	5.63	5.64	5.39	5.49	5.46	5.14	5.26	5.09
Reference material – BOD (mg O2/g)	-	570	536	560	558	592	572	578	600	576	610

	Incubation period (days)
	7		14		21			28
Replicate number	1	2	1	2	1	2	3	1	2	3
Test substance (%)	25	24	34	25	37	36	40	37	36	43
Average degradation (%)	24		29		38			39
Reference material (%)	73	69	72	72	76	73	74	77	74	78
Average degradation (%)	71		72		74			76

Validity criteria fulfilled:

yes

Conclusions:

HPV Xylenols failed to meet the requirements for a pass in this test with a maximum of 39% observed over 28 days. However because of the stringency of the test, this does not necessarily mean that the test substance is not degradable under environmental conditions. The results of the blank tests and reference material show that the tests were conducted correctly and the guidelines followed.

Executive summary:

HPV Xylenols failed to meet the requirements for a pass in this test (³60% degradation relative to the COD value - a maximum of 39% was recorded). However, because of the stringency of the test, this does not necessarily mean that the test substance is not biodegradable under environmental conditions, but indicates that more work would be necessary to establish biodegradability.

 

The guideline requires that the reference material degrades by at least 60% of its ThOD, and that the blank dissolved oxygen loss does not exceed 1.5mg/l after 28 days. Both of these criteria were satisfied for this test; a maximum value of 76% degradation of the sodium acetate was achieved, and the blank DO loss was 0.42mg/l.

 

The ThOD was calculated based on information provided by the client and structures sourced from the Internet.

Description of key information

Study and literature data on cresols and ethylphenols show them to be readily biodegradable.

Study data on xylenols show that 2,3- and 2,4-xylenol are readily biodegradable and the remaining 4 are inherently biodegradable (Pitter, 1976). Based on all available information for cresols, ethyl-phenols and xylenols the substances are described as inherently biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

inherently biodegradable

Additional information

m-, p- and o-Cresol is inherently degradable as shown by a test similar to OECD guideline 302 B with a removal of 95 -96 % of m-cresol after 5 days of incubation. The degradation rates were >15 mg COD/g/h which was the criteria for being readily biodegradable.

Ethyl phenols are considered to be readily biodegradable. The sensitivity of the test system was confirmed as the reference substance sodium benzoate is considered to be readily biodegradable

The xylenols are inherently degradable as shown by a test similar to OECD guideline 302 B with a removal of 89.3 -97.5 % after 5 days of incubation . 2,3 -xylenol and 2,4 -xylenol had degradation rates >15 mg COD/g/h which was the criteria for being readily biodegradable (Pitter 1976)

A recent 301D study (2010) on mixed xylenols showed biodegradation of 39 % in 28 days. However, this test is very stringent and it does not necessarily mean that the substance would not biodegrade under environmental conditions.