N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

January, 22 to February 19, 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1992

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

440/2008/EEC

GLP compliance:

yes (incl. QA statement)

Remarks:

certificate from May 31, 2016

Test material

Test material information

Constituent 1

Reference substance name:

N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide

EC Number:

233-360-4

EC Name:

N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide

Cas Number:

10128-55-9

Molecular formula:

C24H16N2O4S

IUPAC Name:

N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide

Test material form:

solid: particulate/powder

Details on test material:

Name of the test material (as cited in study report): LUMILUX® Yellow CD 397
State of aggregation: white powder

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

Fresh samples of activated sludge are withdrawn on January 22nd, 2018 from the sewage treatment plant Ruhrverband Kläranlage, Sunthelle 6, 57392 Schmallenberg, Germany, which is mainly fed with municipal wastewater. Since it was not necessary, the samples were not washed with mineral medium after the arrival at the laboratory but kept aerobic until use. The sludge was left for settlement for ca. one hour. Subsequently the supernatant was discarded and the concentration of suspended solids was measured in the remaining sludge. The concentration was adjusted to 3.8 g/L and verified by dry mass measurement. The concentration used in the test was 29.6 mg dry mass/liter (7.4 mg dry mass/250 mL).

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

28 mg/L

Based on:

ThOD

Remarks:

test item concentration corresponds to 63 mg ThOD/L

Initial test substance concentration 2

Initial conc.:

100 mg/L

Based on:

ThOD

Remarks:

test item concentration corresponds to 224 mg ThOD/L

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: mineral medium (see table in "any other information on materials and methods")
- Additional substrate: no
- Solubilising agent : no
- Test temperature: 22 °C ± 1°C
- pH (test end): 7.4 - 7.5 (test solutions, inocculum blanks and abiotic control, respectively), 7.9 - 8.0 (process and toxicity control, respectively)
- pH adjusted (start): yes, with 50 g/L H2SO4 to a pH of 7.4 ± 0.2
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: SAPROMAT respirometer (VOITH Inc.)
- vessels per concentration: 2
- Aerobic conditions: created by continuous aeration of the test suspension during the whole test

SAMPLING
- Sampling frequency: continuous measurement of oxygen demand
- Sterility check if applicable: yes

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: yes
- Process control: yes, sodium benzoate
- Toxicity control: yes, sodium benzoate and 100 mg test item/L

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

Preliminary study:

not applicable

Test performance:

No GLP deviations or deviations from the study plan occured.

% Degradation

% Degradation 1

Key result

Parameter:

% degradation (O2 consumption)

Value:

0

St. dev.:

0.9

Sampling time:

28 d

% Degradation 2

Key result

Parameter:

% degradation (O2 consumption)

Value:

-5

St. dev.:

12.8

Sampling time:

28 d

Details on results:

The biodegradation of N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide after 28 days of incubation in the static test was found to be 0% independent from test concentration (actually 0%, SD = 0.9% and -5%, SD = 12.8% in the assays with 100 mg/L and 28 mg/L, respectively). With degradation rates below 10% over the complete test duration in the 100 mg/L assays, no degradation rate within a 10-day-window can be calculated. The biodegradation of the item mixture in the toxicity control was found to be 37% after 14 days of incubation. Thus, the demanded threshold value of 25% is exceeded and the test item N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide can be identified as non-toxic in a ready biodegradability test. The reference item sodium benzoate was degraded to 86% within the first 14 days.

According to the OECD guideline 301F, the test item N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide must be considered as being not readily biodegradable under the test conditions applied.

The test is valid since (1) the difference of extremes of replicate values of the removal of the test item was less than 20 % (i.e. 0 - 6 %) at the end of the 10-day-window and at test end, (2) the percentage degradation of the reference item has exceeded the pass level of 60 % by day 14 and (3) the oxygen uptake of the inoculum blank is < 60 mg/L in 28 days and the mean pH values in all assays were inside the range of 6.0 - 8.5.

BOD5 / COD results

Results with reference substance:

The percentage degradation of the reference item has exceeded the pass level of 60 % by day 14 (i.e. mean % degradation of 85.9 %, SD: 2.5 %).

Any other information on results incl. tables

Table: Cumulated oxygen consumption (in mg O₂/L) after 14 days.

Single and mean values of the parallel test vessels and standard deviation. Test suspension B: 28 mg/L; Test suspension A: 100 mg/L; Procedural control: 100 mg/L; Abiotic control: 100 mg/L; Toxicity control: 200 mg/L.

vessel	Inoculum blank	Test suspension A	Test suspension B	Procedural control	Toxicity control	Abiotic control
1	14	16	12	162	158	0
2	18	19	20	156	165	3
Mean	16	18	16	159	162	2
SD	3	2	6	4	5	2

 

Table: Degradation (in %) after 14 days.

Single and mean values of the parallel test vessels and standard deviation.

vessel		Test suspension A	Test suspension B	Procedural control	Toxicity control	Abiotic control
1		0.0	-6.4	87.7	36.4	0.0
2		1.3	6.4	84.1	38.1	6.7
Mean		0.7	0.0	85.9	37.3	3.3
SD		0.9	9.0	2.5	1.3	4.7

 

Table: Cumulated oxygen consumption (in mg O₂/L) after 28 days.

Single and mean values of the parallel test vessels and standard deviation.Test suspension B: 28 mg/L; Test suspension A: 100 mg/L; Procedural control: 100 mg/L; Abiotic control: 100 mg/L; Toxicity control: 200 mg/L.

vessel	Inoculum blank	Test suspension A	Test suspension B	Procedural control	Toxicity control	Abiotic control
1	24	21	13	180	165	0
2	26	24	24	173	176	4
Mean	25	23	19	177	171	2
SD	1	2	8	5	8	3

 

Table: Degradation (in %) after 28 days.

Single and mean values of the parallel test vessels and standard deviation.

vessel		Test suspension A	Test suspension B	Procedural control	Toxicity control	Abiotic control
1		-1.8	-13.9	93.1	35.8	0.0
2		-0.4	4.2	88.9	38.7	8.9
Mean		-1.1	-4.9	91.0	37.3	4.5
SD		0.9	12.8	3.0	2.0	6.3

 

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

(1) Difference of extremes of replicate values of test item removal were <20% after 10 and 28d, (2) the % degradation of the reference item was < 60 % after 14d and (3) the O2 uptake of inoculum blank was < 60 mg/L in 28d and mean pH values was 6.0 - 8.5.

Interpretation of results:

not readily biodegradable

Conclusions:

The percentage biodegradation of test item Lumilux Yellow CD 397 (N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide) through microorganisms from a municipal sewage treatment plant was investigated in a 28-day Manometric Respirometry Test according to EC method C.4-D and OECD 301 F (1992). Therefore,100 mg/L (ThOD = 224 mg/L) and 28 mg/L (ThOD = 63 mg/L)of the test item were inoculated in mineral medium at22 ± 1°C in darkness and the rate of degradation was monitored by measuring the quantity of oxygen required to maintain a constant gas volume in the respirometer flasks over 28 days.The biodegradation of N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide was 0% for both test concentrations and no degradation rate within a 10-day-window can be calculated since degradation rates remained below 10% over 28 days. All validity criteria were fulfilled and according to OECD 301F, the test item Lumilux Yellow CD 397 (N-[2-(4-oxo-4H-3,1-benzoxazin-2-yl)phenyl]naphthalene-2-sulphonamide) must be considered as being not readily biodegradable.