Registration Dossier

Environmental fate & pathways

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:
2016-03-15 to 2016-04-14
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)
Deviations:
no
Qualifier:
according to
Guideline:
other: ISO International standard 9439 "Water Quality - Evaluation of ultimate Aerobic Biodegradability of organic compounds in aqueous medium - Carbon Dioxide evoluton test (1999)
Deviations:
no
Qualifier:
according to
Guideline:
other: ISO International Standard 10634 "Water quality - Guidance for the preparation and treatment of poorly water-soluble organic compounds for the subsequent evaluation of their biodegradability in an aqueous medium" (1995)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
- Name of test material (as cited in study reports): JNJ-17299412-AAA (T001599)
- Physical state: solid (powder)
- Appearance: white powder
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Batch n°: I15HB2951
- Analytical purity: 100.2% (base titration: >=98.0%)

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Expiration date: 2017-08-09 (re-test date)
- Storage condition: at room temperature
- Stability under storage conditions: Analysis of stability, homogeneity and concentration of the test item under test conditions were not performed as part of this study.

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source: municipal sewage treatment plant receiving predominantly domestic sewage, 'Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands.
- Storage conditions: sludge was kept under continuous aeration until further treatment
- Preparation of inoculum for exposure: Before use, the sludge was allowed to settle (76 minutes) and the supernatant liquid was used as inoculum.
- Pretreatment: no
- Concentration of sludge: the concentration of suspended solids was determined to be 4.0 g/L in the concentrated sludge.
- Water filtered: Tap-water purified by reverse osmosis (Milli-RO) and subsequently passed over activated carbon.
Duration of test (contact time):
28 d
Initial test substance concentrationopen allclose all
Initial conc.:
16.5 mg/L
Based on:
test mat.
Initial conc.:
12 mg/L
Based on:
TOC
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: test water prepared according to test guidelines, analytical grade salts dissolved in tap-water purified by reverse osmosis (Milli-RO) and subsequently passed over activated carbon.
* mineral stock solution A: 8.5 g KH2PO4, 21.75 g K2HPO4, 67.20 g Na2HPO4.12H2O, 0.5 gNH4Cl dissolved in 1 L Milli-Q water, pH 7.4 ± 0.2
* mineral stock solution B: 22.50 g MgSO4.7H2O dissolved in 1 L Milli-Q water
* mineral stock solution C: 36.4 g CaCl2.2H2O dissolved in 1 L Milli-Q water
* mineral stock solution D: 0.25 g FeCl3.6H2O dissolved in 1 L Milli-Q water
* Final test medium: 10 mL of solution A and 1 mL of solutions B, C and D per L of test medium
- Additional substrate: no
- Test temperature: 22.1-22.7°C
- pH: 7.5-7.8, measured prior to testing in each test flask before addition of inoculum, and again in each test flask at the end of the incubation period
- pH adjusted: no data
- Aeration of dilution water: The test solutions were continuously stirred during the test.
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 2-L all-glass brown coloured bottles
- Number of culture flasks/concentration:
* test substance and inoculum: 2 replicates
* inoculum blank: 2 replicates
* positive control: 1 replicate
* toxicity control: 1 replicate
- Method used to create aerobic conditions: A mixture of oxygen (~20%) and nitrogen (~80%) was passed through a bottle, containing 0,5 - 1 L 0,0125 M Ba(OH)2 solution to trap CO2. The synthetic air was sparged through the scrubbing solutions at a rate of ~1-2 bubbles per second ( ~30-100 mL/min). The initial suspension of unspiked test medium and inoculum was aerated with this CO2-free air overnight to purge the system of CO2 prior to testing. This CO2-free air was also used for aeration during the test.
- Measuring equipment: CO2-evolution was determined through titration of the remaining Ba(OH)2 with 0.05 M standardized HCl.
- Details of trap for CO2 and volatile organics if used: Three CO2-absorbers (bottles filled with 100 mL 0.0125 M Ba(OH)2 were connected in series to the exit air line of each test bottle.

SAMPLING
- Sampling frequency: every second or third day during the first 10 days, and thereafter at least every fifth day until the 28th day
- Sampling method: the absorber bottle closest to the incubation system was sampled each time, the second and third bottle were moved one position closer to the system and a new bottle was added at the end
- On the 28th day, pH of test suspensions was measured and 1 mL of concentrated HCl was added to each bottle. Bottles were aerated overnight to drive off CO2 present in the test suspension. The final titration was made on day 29.

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes, two replicates with only inoculum
- Toxicity control: yes, one replicate with test item, reference substance, and inoculum
- Procedure control/positive control: yes, 1 replicate with reference item and inoculum

Reference substance
Reference substance:
acetic acid, sodium salt

Results and discussion

Test performance:
- In the toxicity control more than 25 % degradation occurred within 14 days (38% based on ThCO2). Therefore, the test substance was assumed to be not inhibitory on microbial activity.
- The positive control item was biodegraded by at least 60% (72%) within 14 days.
- The difference of duplicate values for %-degradation of the test item was always less than 20% (≤6%).
- The total CO2 release in the blank at the end of the test did not exceed 40 mg/L (57.7 mg CO2 per 2 litres of medium, corresponding to 28.9 mg CO2/L).
- The Inorganic Carbon content (IC) of the test item (suspension) in the mineral medium at the beginning of the test was less than 5% of the Total Carbon content (TC). Since the test medium was prepared in tap-water purified by reverse osmosis (Milli-RO water (Millipore Corp., Bedford, Mass., USA, carbon levels < 500 ppb)), IC was less than 5% of TC (mainly coming from the test item, 12 mg TOC/L). Since all criteria for validity of the test were met, this study was considered to be valid.
% Degradation
Key result
Parameter:
% degradation (CO2 evolution)
Value:
ca. 4
Sampling time:
28 d
Remarks on result:
other: mean of 2 bottles
Remarks:
Absolute difference between 2 bottles is 6.
Details on results:
The criterion for ready biodegradability (at least 60% biodegradation within a 10-day window) was not met.

BOD5 / COD results

Results with reference substance:
The positive control substance was degraded at least 72 % within 14 days, confirming suitability of the activated sludge.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
not readily biodegradable
Conclusions:
A 28-d ready biodegradability test (OECD 301B, modified sturm test) using unadapted activated sludge from a predominantly domestic waste water treatment plant indicated that JNJ-17299412-AAA (T001599) was not readily biodegradable under the conditions of the test (initial concentration 16.5 mg/L) as the pass level for ready biodegradability was not reached (i.e. biodegradation of at least 60% in a 10-day window period within the 28-day period). the test substance showed only 4 % biodegradation (mean of 2 bottles, based on % ThCO2). The test substance did not inhibit microbial activity at the concentration used in the test and the system was demonstrated to be healthy. The results of the test can be considered reliable without restriction.